Partial protection against enterovirus 71 (EV71) infection in a mouse model immunized with recombinant newcastle disease virus capsids displaying the EV71 VP1 fragment.

Enterovirus 71 (EV71) infection may cause severe neurological complications, particularly in young children. Despite the risks, there are still no commercially available EV71 vaccines. Hence, a candidate vaccine construct, containing recombinant Newcastle disease virus capsids that display an EV71 VP1 fragment (NPt-VP1 1-100) protein, was evaluated in a mouse model of EV71 infection. Previously, it was shown that this protein construct provoked a strong immune response in vaccinated adult rabbits. That study, however, did not address the issue of its effectiveness against EV71 infection in young animals. In the present study, EV71 viral challenge in vaccinated newborn mice resulted in more than 40% increase in survival rate. Significantly, half of the surviving mice fully recovered from their paralysis. Histological analysis of all of the surviving mice revealed a complete clearance of EV71 viral antigens from their brains and spinal cords. In hind limb muscles, the amounts of the antigens detected correlated with the degrees of tissue damage and paralysis. Findings from this study provide evidence that immunization with the NPt-VP1 1-100 immunogen in a newborn mouse model confers partial protection against EV71 infection, and also highlights the importance of NPt-VP1 1-100 as a possible candidate vaccine for protection against EV71 infections

Immunization with recombinant enterovirus 71 viral capsid protein 1 fragment stimulated antibody responses in hamsters

Enterovirus 71 (EV71) causes severe neurological diseases resulting in high mortality in young children worldwide. Development of an effective vaccine against EV71 infection is hampered by the lack of appropriate animal models for efficacy testing of candidate vaccines. Previously, we have successfully tested the immunogenicity and protectiveness of a candidate EV71 vaccine, containing recombinant Newcastle disease virus capsids that display an EV71 VP1 fragment (NPt-VP1(1-100)) protein, in a mouse model of EV71 infection. A drawback of this system is its limited window of EV71 susceptibility period, 2 weeks after birth, leading to restricted options in the evaluation of optimal dosing regimens. To address this issue, we have assessed the NPt-VP1(1-100) candidate vaccine in a hamster system, which offers a 4-week susceptibility period to EV71 infection. Results obtained showed that the NPt-VP1(1-100) candidate vaccine stimulated excellent humoral immune response in the hamsters. Despite the high level of antibody production, they failed to neutralize EV71 viruses or protect vaccinated hamsters in viral challenge studies. Nevertheless, these findings have contributed towards a better understanding of the NPt-VP1(1-100) recombinant protein as a candidate vaccine in an alternative animal model system

A monoclonal antibody to ameliorate central nervous system infection and improve survival in a murine model of human Enterovirus-A71 encephalomyelitis

Background: Enterovirus A71 (EV-A71) encephalomyelitis is an often fatal disease for which there is no specific treatment available. Passive immunization with a specific monoclonal antibody to EV-A71 was used on a murine model of EV-A71 encephalomyelitis to evaluate its therapeutic effectiveness before and after established central nervous system (CNS) infection. Methods: Mice were intraperitoneally-infected with a mouse-adapted EV-A71 strain and treated with a dose of monoclonal antibody (MAb) daily for 3 days on day 1, 2 and 3 post-infection or for 3 days on 3, 4 and 5 post-infection. Treatment effectiveness was evaluated by signs of infection and survival rate. Histopathology and qPCR analyses were performed on mice sacrificed a day after completing treatment. Results: In mock-treated mice, CNS infection was established from day 3 post-infection. All mice treated before established CNS infection, survived and recovered completely without CNS infection. All mice treated after established CNS infection survived with mild paralysis, and viral load and antigens/RNA at day 6 post-infection were significantly reduced. Conclusions: Passive immunization with our MAb could prevent CNS infection in mice if given early before the establishment of CNS infection. It could also ameliorate established CNS infection if optimal and repeated doses were given

Pathological findings in a mouse model for Coxsackievirus A16 infection

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

Neuronal transcriptomic responses to Japanese encephalitis virus infection with a special focus on chemokine CXCL11 and pattern recognition receptors RIG-1 and MDA5

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 < 0.05) were found at 48 hours post-infection (hpi) and 60 hpi, respectively. The genes were mainly involved in anti-microbial responses, cell signalling, cellular function and maintenance, and cell death and survival. Among the most highly upregulated genes (≥ 10 folds, P < 0.05) were chemokines CCL5, CXCL11, IL8 and CXCL10. The upregulation and expression of CXCL11 were confirmed by qRT-PCR and immunofluorescence. Pathogen recognition receptors retinoic acid-inducible gene-1 (RIG-1) and melanoma differentiation-associated protein 5 (MDA5) were also upregulated. Our results strongly suggest that neuronal cells play a significant role in immunity against JEV. CXCL11, RIG-1 and MDA5 and other cytokines may be important in neuropathogenesi

Lethal Nipah Virus Infection Induces Rapid Overexpression of CXCL10

Nipah virus (NiV) is a recently emerged zoonotic Paramyxovirus that causes regular outbreaks in East Asia with mortality rate exceeding 75%. Major cellular targets of NiV infection are endothelial cells and neurons. To better understand virus-host interaction, we analyzed the transcriptome profile of NiV infection in primary human umbilical vein endothelial cells. We further assessed some of the obtained results by in vitro and in vivo methods in a hamster model and in brain samples from NiV-infected patients. We found that NiV infection strongly induces genes involved in interferon response in endothelial cells. Among the top ten upregulated genes, we identified the chemokine CXCL10 (interferon-induced protein 10, IP-10), an important chemoattractant involved in the generation of inflammatory immune response and neurotoxicity. In NiV-infected hamsters, which develop pathology similar to what is seen in humans, expression of CXCL10 mRNA was induced in different organs with kinetics that followed NiV replication. Finally, we showed intense staining for CXCL10 in the brain of patients who succumbed to lethal NiV infection during the outbreak in Malaysia, confirming induction of this chemokine in fatal human infections. This study sheds new light on NiV pathogenesis, indicating the role of CXCL10 during the course of infection and suggests that this chemokine may serve as a potential new marker for lethal NiV encephalitis

Idiopathic ileal volvulus with multiple concomitant infections in a starving man

Background: Small bowel volvulus is defined as the torsion of the small intestine, potentially leading to bowel obstruction, gangrene and perforation. It is a rare condition, especially in adults. Case presentation: A 30-year-old man was retrieved from the jungle with severe weight loss and abdominal symptoms. He succumbed to death despite 22 days of intensive medical treatment. An autopsy revealed a ruptured gangrenous ileal volvulus with peritonitis and subdiaphragmatic abscess. Further laboratory analysis detected systemic Candida tropicalis and intestinal gramnegative bacterial sepsis, systemic Zika virus viremia, leptospirosis complicating rhabdomyolysis and disseminated intravascular coagulopathy, Type I Herpes Simplex virus infection of the tongue and upper gastrointestinal tract. The cause of death was the ruptured ileal volvulus, complicated with upper gastrointestinal bleeding due to Herpes simplex virus esophagitis in a malnourished patient with resolving leptospirosis and underlying Zika virus co-infection. Conclusion: Rare clinical scenarios of adult-onset intestinal volvulus with concomitant multiple infections precludes clinical diagnosis and early treatment, leading to devastating consequences of clinical outcome. The positive clinical and postmortem correlation is a good learning lesson in many disciplines of medicine and science

Sarcocystis nesbitti causes acute, relapsing febrile myositis with a high attack rate: Description of a large outbreak of muscular sarcocystosis in Pangkor Island, Malaysia, 2012

Background: From the 17th to 19th January 2012, a group of 92 college students and teachers attended a retreat in a hotel located on Pangkor Island, off the west coast of Peninsular Malaysia. Following the onset of symptoms in many participants who presented to our institute, an investigation was undertaken which ultimately identified Sarcocystis nesbitti as the cause of this outbreak. Methodology/Principal Findings: All retreat participants were identified, and clinical and epidemiological information was obtained via clinical review and self-reported answers to a structured questionnaire. Laboratory, imaging and muscle biopsy results were evaluated and possible sources of exposure, in particular water supply, were investigated. At an average of 9-11 days upon return from the retreat, 89 (97) of the participants became ill. A vast majority of 94 had fever with 57 of these persons experiencing relapsing fever. Myalgia was present in 91 of patients. Facial swelling from myositis of jaw muscles occurred in 9 (10) patients. The median duration of symptoms was 17 days (IQR 7 to 30 days; range 3 to 112). Out of 4 muscle biopsies, sarcocysts were identified in 3. S. nesbitti was identified by PCR in 3 of the 4 biopsies including one biopsy without observed sarcocyst. Non-Malaysians had a median duration of symptoms longer than that of Malaysians (27.5 days vs. 14 days, p = 0.001) and were more likely to experience moderate or severe myalgia compared to mild myalgia (83.3 vs. 40.0, p = 0.002). Conclusions/Significance: The similarity of the symptoms and clustered time of onset suggests that all affected persons had muscular sarcocystosis. This is the largest human outbreak of sarcocystosis ever reported, with the specific Sarcocystis species identified. The largely non-specific clinical features of this illness suggest that S. nesbitti may be an under diagnosed infection in the tropics

Development of live attenuated Enterovirus 71 vaccine strains that confer protection against lethal challenge in mice

Besides causing mild hand, foot and mouth infections, Enterovirus A71 (EV-A71) is associated with neurological complications and fatality. With concerns about rising EV-A71 virulence, there is an urgency for more efective vaccines. The live attenuated vaccine (LAV) is a more valuable vaccine as it can elicit both humoral and cellular immune responses. A miRNA-based vaccine strain (pIY) carrying let-7a and miR-124a target genes in the EV-A71 genome which has a partial deletion in the 5′NTR (∆11bp) and G64R mutation (3Dp°l ) was designed. The viral RNA copy number and viral titers of the pIY strain were signifcantly lower in SHSY-5Y cells that expressed both let-7a and miR-124a. Inhibition of the cognate miRNAs expressed in RD and SHSY-5Y cells demonstrated de-repression of viral mRNA translation. A previously constructed multiply mutated strain, MMS and the pIY vaccine strain were assessed in their ability to protect 4-week old mice from hind limb paralysis. The MMS showed higher amounts of IFN-γ ex vivo than the pIY vaccine strain. There was absence of EV-A71 antigen in the skeletal muscles and spinal cord micrographs of mice vaccinated with the MMS and pIY strains. The MMS and pIY strains are promising LAV candidates developed against severe EV-A71 infections