Prospective Investigation of Oxidative Stress in West Nile Virus Infection (PROWENI)

The investigator hypothesizes that oxidative stress responses to West Nile virus infection in the central nervous system determine the severity of infection and the long-term neurological, neuropsychological and functional sequelae of West Nile Neuroinvasive Disease.

Study Overview

• Status: Withdrawn
• Conditions: Oxidative Stress; West Nile Virus; West Nile Fever; West Nile Fever Encephalitis; West Nile Fever Meningitis; West Nile Meningitis; West Nile Virus Meningoencephalitis; West Nile Fever Meningoencephalitis; West Nile Fever With Neurologic Manifestation

• Study Type: Observational

Contacts and Locations

Study Locations

• Romania
  • Bucharest, Romania
    • Victor Babes Hospital

Participation Criteria

Eligibility Criteria

• Ages Eligible for Study: 14 years and older (Adult, Older Adult)
• Accepts Healthy Volunteers: No
• Genders Eligible for Study: All

• Sampling Method: Non-Probability Sample

Study Population

The study population will be recruited from VBH emergency and outpatient departments in Bucharest, Romania, from the onset of the WNV transmission season in 2020 until November 2023. The incidence of WNV and WNND in South-Eastern Romania was 0.5 per 100,000 inhabitants in 2016, and has been increasing steadily 35. Romania reported 277 WNND cases with 43 deaths in 2018, and 66 WNND cases with 8 deaths in 2019 3. VBH is a tertiary care facility that serves a population of 3 million people (15% of Romania's population). Its catchment area covers more than 80% of the territories in Romania that reported WNV transmission and has in recent years been the largest single medical center to care for WNV infected patients.

Description

Inclusion Criteria:

• Willing and able to provide written informed consent. If the clinical condition of the patient does not permit giving consent, informed consent will be obtained from the next of kin.
• Age 18 years or older
• for active cases: positive anti-WNV IGM antibodies in serum (or IgG in CSF if applicable)
• for active cases: presentation within (maximum) 7days of symptom onset
• for healthy controls: anti-WNV antibody naive (IgM and IgG in serum). The group of healthy controls will be selected to have an age similar distribution to the cases.

Exclusion Criteria:

• Evidence of active systemic infection in 3 months prior to recruitment
• Evidence of systemic inflammatory illness
• Clinical signs of neurodegenerative or neurologic disease other than WNND
• Pregnancy
• Active malignancy
• History of drug abuse

Study Plan

How is the study designed?

Design Details

• Observational Models: Case-Control
• Time Perspectives: Prospective

Number of groups / cohorts

3

Cohorts and Interventions

Group / Cohort
WNND; 40 subject with West-Nile Neuroinvasive Disease will be recruited
WNF; 40 subject with West-Nile Fever will be recruited
controls; 20 control will be recruited. These controls will be aged matched to the cases.

What is the study measuring?

Primary Outcome Measures

Outcome Measure	Measure Description	Time Frame
Measure the redox status	A multiparameter indexes of oxidative stress will be calculated to measure and summarize the redox status in cases and age matched controls. Association between the redox status and clinical, neuropsychological and radiological outcomes will be investigated. We will also examine the relative sensitivity of separate biomarkers of oxidative stress and autophagy as clinical predictors of WNV infection severity.	at recruitment, 10 days post-symptom onset and 20 days post symtom onset.
Assessment of Neurologic deficits	As part of the descriptive analysis of biomarkers of disease severity and to study to neurologic sequelae of WNV infection. Will be assessed: specifically assessments of cranial nerves II- XII, motor strength in upper and lower extremities, sensory testing for pinprick and vibration, deep tendon reflexes, gait, coordination, and movement abnormalities	At recruitment, month 3 and month 12
Neuropsychologic performance	Study the neuropsychologic sequelae of WNV infection during a 12-month followup period in following key domains: Attention, Memory, Executive Function, Emotion & Social Cognition, Psychomotor Speed	20 days post-symtom onset, month 3 and month 12
Longitudinal assessment of functional status Study the neurologic and neuropsychologic sequelae of WNV infection during a 12-month followup period.	ECOG/WHO PS during a 12-month followup.	at recruitment, month 3 and month 12
MRI abnormalities	Part of descriptive analysis of clinical markers of disease severity	at recruitment, month 3 and month 12
Brain iron content	Part of the descriptive analysis of clinical markers of disease severity: Qualitative analysis per neuroanatomical region by iron-sensitive MRI sequence (SWI)	at recruitment, month 3 and month 12
Ophthalmological abnormalities	As part of the descriptive analysis of clinical markers of disease severity ophthalmologic abnormalities will be assessed by slit lamp examination	at recruitment, with a follow-up of clinically indicated.
serum S100b concentration	As part of the descriptive analysis of clinical markers of disease severity S100b concentration will be measured to asses the Blood-Brain barrier integrity	at recruitment, 10 days post symptom onset and 20 days post symptom onset
serum NSE concentration	As part of the descriptive analysis of clinical markers of disease severity NSE concentration will be measured to asses the Blood-Brain barrier integrity	at recruitment, 10 days post symptom onset and 20 days post symptom onset

Secondary Outcome Measures

Outcome Measure	Measure Description	Time Frame
Analysis of laboratory performance characteristics (e.g. sensitivity) of WNV-specific RT-PCR and viral isolation in clinical samples, compared to composite diagnosis of WNV infection	Confirmed WNV infection is defined as one or more of following criteria: Seroconversion of anti-WNV IgG antibodies (Indirect immunofluorescence testing (IIFT) Flavivirus 2, Euroimmun®, Lübeck, Germany) in convalescent serum (20 days after symptom onset) (all participants); 4-fold increase of anti-WNV IgG antibodies in convalescent serum, compared to the screening sample (IIFT).; Anti-WNV IgM antibody detection (ELISA) in CSF* (at VBH) (*CSF sampled only when clinically indicated).; Positive WNV-specific RT-PCR result in serum, urine or CSF in samples obtained at screening.; Virus isolation by outgrowth assay in samples obtained at screening	20 days
Description of molecular epidemiology of infecting WNV strain(s) and viral outgrowth diagnostic performance.	The time frame for WNV detection after symptom onset by real-time reverse transcriptase polymerase chain reaction (RT-PCR) or virus isolation by outgrowth from blood or CSF is limited by the fast decline of circulating virus. Because the WNV RNA detection window in urine can be longer (up to 14 days), RT-PCR will be performed on all clinical samples at recruitment. The target sequences are a conserved region in the 5'UTR region and part of the capsid gene of WNV and detects both lineage 1 and 2 WNV. WNV has been successfully isolated from urine, in approx. 40% of samples obtained within 8 days of symptom onset. WNV isolation will be attempted in low-passage Vero E6 cells and BHK21 cells from urine samples with a high WNV RNA load (Cycle threshold (Ct)- values <30).	20 days
Identification of potential genetic signatures that correlate with virulence (neuro-invasion and morbidity) in our cohort.	Whole genome sequencing (WGS) will be performed on a subset of RT-PCR positive serum, urine and CSF samples. This will allow us to investigate the temporal and spatial compartmentalization of the West Nile lineages in Romania as well as expand our understanding of genetic compartmentalization within hosts (urine, serum, CSF). In addition, the obtained isolates will be sequenced (from the viral outgrowth assay) to assess potential adaptations the virus undergoes during isolation. Finally, in a genome wide association study, the obtained viral sequences will be compared with the prospectively collected clinical data, to identify potential genetic signatures that correlate with virulence, neuroinvasion and morbidity. Our data will be compared to existing data on genetic determinants of virulence such as the presence of a glycosylation site in the E protein, substitutions in non-structural proteins 3, 4B or 5, or variation in the 3' noncoding region.	20 days

Collaborators and Investigators

• Sponsor: Institute of Tropical Medicine, Belgium
• Collaborators: Universiteit Antwerpen; Victor Babes Hospital, Bucharest, Romania
• Investigators: Principal Investigator: Koen Vercauteren, Institute Of Tropical Medicine Antwerp; Principal Investigator: Nina Hermans, Universiteit Antwerpen; Principal Investigator: Corneliu Popescu, Victor Babes Hospital, Bucharest

Study record dates

Study Major Dates

• Study Start (Anticipated): September 1, 2020
• Primary Completion (Anticipated): October 1, 2023
• Study Completion (Anticipated): December 31, 2023

Study Registration Dates

• First Submitted: April 9, 2020
• First Submitted That Met QC Criteria: April 28, 2020
• First Posted (Actual): May 1, 2020

Study Record Updates

• Last Update Posted (Actual): February 10, 2021
• Last Update Submitted That Met QC Criteria: February 9, 2021
• Last Verified: February 1, 2021

More Information

Terms related to this study

• Keywords: Genome sequencing; composite diagnosis
• Additional Relevant MeSH Terms: Brain Diseases; Central Nervous System Diseases; Nervous System Diseases; RNA Virus Infections; Infections; Encephalitis, Arbovirus; Encephalitis, Viral; Central Nervous System Viral Diseases; Central Nervous System Infections; Infectious Encephalitis; Arbovirus Infections; Vector Borne Diseases; Flavivirus Infections; Flaviviridae Infections; Body Temperature Changes; Encephalitis; Virus Diseases; Fever; Meningitis; Neurologic Manifestations; Meningoencephalitis; West Nile Fever
• Other Study ID Numbers: ITM202005

Drug and device information, study documents

• Studies a U.S. FDA-regulated drug product: No
• Studies a U.S. FDA-regulated device product: No