Development of a High-throughput Sequencing Platform for Detection of Viral Encephalitis Pathogens Based on Amplicon Sequencing

Objective Viral encephalitis is an infectious disease severely affecting human health.It is caused by a wide variety of viral pathogens,including herpes viruses,flaviviruses,enteroviruses,and other viruses.The laboratory diagnosis of viral encephalitis is a worldwide challenge.Recently,high-throughput sequencing technology has provided new tools for diagnosing central nervous system infections.Thus,In this study,we established a multipathogen detection platform for viral encephalitis based on amplicon sequencing.Methods We designed nine pairs of specific polymerase chain reaction(PCR)primers for the 12 viruses by reviewing the relevant literature.The detection ability of the primers was verified by software simulation and the detection of known positive samples.Amplicon sequencing was used to validate the samples,and consistency was compared with Sanger sequencing.Results The results showed that the target sequences of various pathogens were obtained at a coverage depth level greater than 20×,and the sequence lengths were consistent with the sizes of the predicted amplicons.The sequences were verified using the National Center for Biotechnology Information BLAST,and all results were consistent with the results of Sanger sequencing.Conclusion Amplicon-based high-throughput sequencing technology is feasible as a supplementary method for the pathogenic detection of viral encephalitis.It is also a useful tool for the high-volume screening of clinical samples.

The Diagnostic Value of Oligoclonal Band Detection in Viral Encephalitis

Objective: This study aims to explore the differences in cerebrospinal fluid oligoclonal band (CSF-OCB) expression among different age groups in viral encephalitis and its reference value for diagnosis. Methods: Forty-two patients with viral encephalitis were divided into two groups: 25 adults and 17 children. The presence of oligoclonal bands in the cerebrospinal fluid (CSF) was detected using polyacrylamide gel electrophoresis, and CSF routine analysis was conducted for comparative analysis. Results: The CSF-OCB positivity rate was higher in the adult group (48%) compared with the pediatric group (11.76%), with a statistically significant difference (P Conclusion: 1) The expression of CSF-OCB positivity in patients with viral encephalitis is age-related, with higher positivity rates observed in adults compared to children. 2) Although CSF oligoclonal band detection is not a specific diagnostic marker for viral encephalitis in adults, it still holds certain reference value.

Analysis of the Effectiveness of Targeted Nursing in Children with Severe Viral Encephalitis Complicated with Respiratory Failure

Objective:To explore and analyze the effectiveness of targeted nursing in children with severe viral encephalitis complicated with respiratory failure.Methods:From April 2021 to April 2023,74 children with severe viral encephalitis complicated with respiratory failure admitted to the Department of Pediatrics of our hospital were selected as the research objects and divided into the target group(n=37)and the reference group(n=37).Targeted nursing was given to the target group,while general nursing was given to the reference group.Physical rehabilitation,motor scores,and lung function were compared between the groups.Results:The recovery time of limb abnormalities,convulsions,abnormal consciousness,and cranial nerve disorders in the target group was significantly better than that in the reference group(P<0.05).The motor function,joint activity and pain,sensory function,and total score of the target group were significantly lower than those of the reference group(P<0.05).After the intervention,lung function indicators including VC(vital capacity),FVC(forced vital capacity),and MVV(maximum voluntary ventilation)of the target group were better than those of the reference group(P<0.05).Conclusion:Targeted nursing can shorten the recovery period of severe viral encephalitis complicated with respiratory failure in children,speed up the recovery of motor function,and improve lung function.This nursing model has a significant application effect in children with severe viral encephalitis complicated with respiratory failure.

Advances in viral encephalitis:Viral transmission,host immunity,and experimental animal models

Viral infections have led to many public health crises and pandemics in the last few centuries.Neurotropic virus infection-induced viral encephalitis(VE),especially the symptomatic inflammation of the meninges and brain parenchyma,has attracted growing attention due to its high mortality and disability rates.Understanding the infectious routes of neurotropic viruses and the mechanism underlying the host immune response is critical to reduce viral spread and improve antiviral therapy outcomes.In this review,we summarize the common categories of neurotropic viruses,viral transmission routes in the body,host immune responses,and experimental animal models used for VE study to gain a deeper understanding of recent progress in the pathogenic and immunological mechanisms under neurotropic viral infection.This review should provide valuable resources and perspectives on how to cope with pandemic infections.

Shuanghuanglian injection downregulates nuclear factor-kappa B expression in mice with viral encephalitis

A mouse model of viral encephalitis was induced by intracranial injection of a Coxsackie virus B3 suspension. Quantitative real-time reverse transcription-PCR and western blot assay were applied to detect mRNA and protein expression of intelectin-2 and nuclear factor-kappa B in the viral encephalitis and control groups. Nuclear factor-kappa B and intelectin-2 mRNA and protein expression were significantly increased in mice with viral encephalitis. After intraperitoneal injection of Shuanghuanglian at a dose of 1.5 mg/kg for 5 successive days, intelectin-2 and nuclear factor-kappa B protein and mRNA expression were significantly decreased. To elucidate the relationship between intelectin-2 and nuclear factor-kappa B, mice with viral encephalitis were administered an intracerebral injection of 107 pfu recombinant lentivirus expressing intelectin shRNA. Both protein and mRNA levels of intelectin and nuclear factor-kappa B in brain tissue of mice were significantly decreased. Experimental findings suggest that Shuanghuanglian injection may downregulate nuclear factor-kappa B production via suppression of intelectin production, thus inhibiting inflammation associated with viral encephalitis.

Inhibitory effects of Shuanghuanglian injection on nuclear factor-kappa B expression in mice with viral encephalitis in a time-and dose-dependent manner

Previous studies have confirmed that the anti-virus effects of Shuanghuanglian injection may be associated with nuclear factor-kappa B activity. This study observed nuclear factor-kappa B expression in mice with viral encephalitis, and showed significant decreases in nuclear factor-kappa B protein and mRNA levels following Shuanghuanglian injection. The inhibitory effect was more significant with prolonged intervention duration and increased treatment dose. These findings verify that Shuanghuanglian injection plays a therapeutic role in viral encephalitis by reducing expression of nuclear factor-kappa B in a time- and dose-dependent manner.

Clinical Efficacy and Application Progress of Reduning Injection in Children with Viral Encephalitis

Viral encephalitis in children caused by multiple viral infections,as a common central nervous system disease in pediatrics,has a high incidence in China.Its clinical symptoms usually include infection,poisoning,intracranial hypertension,and disturbance of consciousness.In severe cases,it can even endanger the lives of patient children.At present,the clinical treatment of the viral encephalitis mainly uses traditional Chinese medicine preparations such as Reduning Injection.The efficacy is significant.It can not only relieve the uncomfortable symptoms in a short time,but also has low incidence of adverse reactions.In this study,the factors that cause viral encephalitis in children were analyzed,and the efficacy,pharmacological action and clinical application of Reduning Injection were comprehensively elaborated,to provide a reference for clinical doctors to rationally treat viral encephalitis in children.

Alterative application of five anticonvulsants according to the half life for the treatment of status epilepticus in children with severe viral encephalitis

BACKGROUND： Traditional subhibernation therapy may easily cause complications, such as respiratory depression and hyportension because of application of chlorpromazine hydrochloride and promethazine in a large dosage. OBJECTIVE： To observe therapeutic effect of modified subhibernation therapy （alterative application of five anticonvulsants according to the half life） on status epilepticus in children with severe viral encephalitis （VE）. DESIGN： Contrast observation. SETTING： Department of Pediatrics, the First Hospital of Jilin University. PARTICIPANTS： The participants in present study were 96 patients with severe viral encephalitis including 52 boys and 44 girls who received treatment in the Department of Pediatrics, the First Hospital of Jilin University from February 2000 to March 2006. All children met the diagnostic criteria of Zhufutong Practice Pediatrics （the seventh edition）. Two weeks ago, they ever got upper respiratory infection or enteronitis and so on before the onset, spirit abnormal, behavior disorder, limbs act disorder, vomit, headache, convulsion, nervous system masculine signs such as limbs act disord, autonomic nerve damage manifestation, brain nerve palsy, dysreflexia, meningeal irritation sign, cerebrospinal fluid and electroencephalography （EEG） abnormity. All parents provided the confirmed consent. The patients were randomly divided into control group （n =40） and experimental group （n =56）. METHODS： Patients in the control group received anticonvulsion, ice compress and routine treatment. The convulsion was treated with five drugs： 0.5 mg/kg wintermin and phenergan, respectively, 100 g/L chlorpromazine hydrochloride （0.5 mL/kg）, 5 mg/kg luminal, 0.3 mg/kg ansiolin. When convulsion attacked, those five drugs were given alternatively; however, those were not given if the convulsion did not attack. Children in the experimental group were treated with improved subhibernation therapy based on routine treatment. The dosages of anticonvulsants were as the same as those in the control group. Based on the half life, every drug was alternated every 4-6 hours. In addition, anticonvulsants administrated for 2 successive days whether tic attacked or not. Then the hypnotic was removed gradually. MAIN OUTCOME MEASURES： Therapeutic efficacy, time of disappeared clinical symptoms and physical sign, and security of administration. RESULTS： All the 96 patients were involved in the final analysis. ① Total effective rate and reliability： Total effective rate was higher in the treatment groups than the control group （χ2=5.871 7, P 〈 0.05）. All patients did not have respiratory depression and side effects. ② Time of disappeared clinical symptoms and physical sign： Recovery time of convulsion, fever, headache and vomit was shorter in the treatment group than that in the control group, and there was significant difference （t =17.612 1-34.330 7, P 〈 0.05）; in addition, symptoms of status epilepticus were relieved obviously. Meanwhile, recovery time of paralysis, coma and anepia was shorter in the treatment group than that in the control group, and there was significant difference （t =10.660 8-24.700 8, P 〈 0.05）. CONCLUSION： Therapeutic effect of improved subhibernation therapy on status epilepticus induced by severe viral encephalitis is positively and safer.

The roles of microglia in viral encephalitis:from sensome to therapeutic targeting

Viral encephalitis is a devastating disease with high mortality,and survivors often suffer from severe neurological complications.Microglia are innate immune cells of the central nervous system(CNS)parenchyma whose turnover is reliant on local proliferation.Microglia express a diverse range of proteins,which allows them to continuously sense the environment and quickly react to changes.Under inflammatory conditions such as CNS viral infection,microglia promote innate and adaptive immune responses to protect the host.However,during viral infection,a dysregulated microglia-T-cell interplay may result in altered phagocytosis of neuronal synapses by microglia that causes neurocognitive impairment.In this review,we summarize the current knowledge on the role of microglia in viral encephalitis,propose questions to be answered in the future and suggest possible therapeutic targets.

Differential transcriptomic landscapes of multiple organs from SARS-CoV-2 early infected rhesus macaques

SARS-CoV-2 infection causes complicated clinical manifestations with variable multi-organ injuries,how-ever,the underlying mechanism,in particular immune responses in different organs,remains elusive.In this study,comprehensive transcriptomic alterations of 14 tissues from rhesus macaque infected with SARS-CoV-2 were analyzed.Compared to normal controls,SARS-CoV-2 infection resulted in dysregulation of genes involving diverse functions in various examined tissues/organs,with drastic transcriptomic changes in cerebral cortex and right ventricle.Intriguingly,cerebral cortex exhibited a hyperinflammatory state evidenced by sig-nificant upregulation of inflammation response-related genes.Meanwhile,expressions of coagulation,angio-genesis and fibrosis factors were also up-regulated in cerebral cortex.Based on our findings,neuropilin 1(NRP1),a receptor of SARS-CoV-2,was significantly elevated in cerebral cortex post infection,accompanied by active immune response releasing inflammatory factors and signal transmission among tissues,which enhanced infection of the central nervous system(CNS)in a positive feedback way,leading to viral encephalitis.Overall,our study depicts a multi-tissue/organ tran-scriptomic landscapes of rhesus macaque with early infection of SARS-CoV-2,and provides important insights into the mechanistic basis for COVID-19-asso-ciated clinical complications.