Research priorities for Nipah Virus Disease in India ICMR-NIV mandate: Research on viruses of Public health Importance -Laboratory based research/-Diagnostic preparedness -Training & capacity building for NiV diagnosis in India -Point of care tests Dr. Pragya D Yadav, M.Sc., PhD, Scientist 'E’, ICMR –National Institute of Virology, Sus Road, Pashan, Pune, India
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Research priorities for Nipah Virus Disease in India · Research priorities for Nipah Virus Disease in India ICMR-NIV mandate: Research on viruses of Public health Importance-Laboratory
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Research priorities for Nipah Virus Disease in India
ICMR-NIV mandate: Research on viruses of Public health Importance
-Laboratory based research/-Diagnostic preparedness
-Training & capacity building for NiV diagnosis in India
-Point of care tests
Dr. Pragya D Yadav, M.Sc., PhD,Scientist 'E’, ICMR –National Institute of
Virology, Sus Road, Pashan, Pune, India
Conventional Nipah virus diagnostic methods
Molecular assays– Real-time RT-PCR
– Conventional RT-PCR and Sanger sequencing
Serological assays– Anti Nipah Human IgM [Sample screening]
– Anti Nipah Human IgG ELISA [serosurvey or diagnosis]
– Anti Nipah Bat IgG ELISA [Bat serosurvey]
– Anti Nipah Swine IgG ELISA [Swine serosurvey]
Virus isolation and characterisation
Antigen detection– Immunofluorescence
– Immunohistochemistry
Commercial assays : None
• Require Containment Laboratory for handling specimen2
Commonly used
Diagnosticmethods
3
Day Post illness
NiV RNA positive/samples
(%positivity)
IgM positive/Total samples
(%positivity)
IgG positive/Total samples
(%positivity)
Day 2 2/2 (100) 1/2 (50) 0/2 (0)
Day 3 3/3 (100) 1/3 (33) 1/3 (33)
Day 5 5/5 (100) 4/5 (80) 0/5 (0)
Day 6 5/5 (100) 3/5 (60) 2/5 (40)
Day 7 2/2 (100) 2/2 (100) 0/2 (0)
Day 9 1/1 (100) 1/1 (100) 1/1 (100)
Detection of Nipah viral RNA and antibody of patients of Kerala Outbreak till 9th POD (n=18)
Detection of Nipah antibody of patients of Malaysia Outbreak, 1999
4
Day Post illness
IgM % positivity
IgG % positivity
Day 1 44%-50% 27%-31%
Day 4 60%-71% 7%-10%
Day 12 100% 40%-44%
Day 17 100% 100%
3 months 95% 100%
Total Number of patients tested: 176Reference: Ramasundrum, et al., Neurol J Southeast Asia, 2000
Cumulative seropositive
response
IgM seropositivity from Day of
illness
IgG seropositivity from Day of illness
25% Day 5 Day 8
50% Day 11 Day 18
75% Day 15 Day 34
95% Day 50 Day 193
Cumulative IgM seropositive response
Cumulative IgG seropositive response
NiV Seropositivity in patient [Bangladesh and India Outbreaks]
5
Day Post illness
IgM positive/Total
samples (%positivity)
IgG positive/Total
samples (%positivity)
Day 1-4 3/5 (60%) 1/5 (20%)
Day 5-10 22/24 (92%) 10/24 (42%)
Day 11-14 9/9 (100%) 6/9 (67%)
Day 15-30 4/4 (100%) 4/4 (100%)
Day 45-60 3/4 (75%) 4/4 (100%)
>1.5 years 0/4 (0%) 4/4 (100%)
Day Post illness
IgM positive/Total
samples(% positivity)
IgG positive/Total
samples(% positivity)
Day 1-3 2/5 (40%) 3/5 (60%)
Day 5 2/2 (100%) 2/2 (100%)
Day 6 1/1 (100%) 1/1 (100%)
Day 7 0/2 (0%) 0/2 (0%)
Day 8 1/2 (50%) 1/2 (50%)
Day 9 1/2 (50%) 1/2 (50%)
Day 10 2/3 (67%) 2/3 (67%)
Bangladesh Outbreak, 1999Total Number of patients tested: 50
Hossain, et al., Clinical Infectious Diseases, 2008
India Outbreak, 2001Total Number of patients tested: 17Chadha, et al., Emerging Infectious
Diseases, 2006
NiV isolations in different Nipah outbreaks
POD NiV isolation from TS*
/Total samples (% positivity)
NiV isolation from NS*
/Total samples
(% positivity)
NiV isolation from urine/Total
samples(% positivity)
1 1/1 (100) 0/1 (0) 0/1 (0)
2 3/5 (60) 1/5 (20) 2/5 (40)
3 0/2 (0) 0/2 (0) 1/2 (50)
4 1/5 (20) 0/5 (0) 1/5 (20)
5 1/4 (25) 0/4 (0) 0/4 (0)
7 0/3 (0) 0/3 (0) 0/3 (0)
Reference: Chua, et al., Journal of Infection, 2001
Malaysia Outbreak, 1999
POD NiV isolation from TS*/Total
samples (% positivity)
NiV isolation from CSF*/Total
samples(% positivity)
3 1/1 (100) 0/1 (0)
5 0/0 0/1 (0)
6 1/2 (50) 0/2 (0)
8 0/1 (0) 0/1 (0)
9 0/1 (0) 0/1 (0)
Reference: Rahman, et al., Vector-Borne and Zoonotic Diseases, 2012
Two NiV isolates were obtained from CSF of two index death cases on Day 4 and 16
No isolate was obtained from 18 serum samples and 6 urine samples from NiV-positive samples
Siliguri, India Outbreak, 2001
Nipah Virus isolation from Kerala Outbreak [2018] in VeroCCL81 cells
Only one NiV isolate was obtained from throat swab (POD:3) Total number of samples 23 processed of 9 NiV positive cases (7-fatal, 2-survival)
Control Vero CC81 cells Infected cells at 1st PID Infected cells at 2nd PID
Cytopathic effects in throat swab Infected VeroCCL81 cells at 2nd PID
Commercial Nipah diagnostic assays (?)
No commercial Serodiagnostic assays are available for detection of Anti Nipah Human IgM antibodyAnti Nipah Human IgG antibodyAnti Nipah Bat IgG antibodyAnti Nipah Pig IgG antibody
Krishgen Biosystems, Mumbai, India: Purchased ELISA kit for Nipah Human IgM and IgG detection but failed to detect positive controls
Pen-side strip test for the detection of Nipah virus in swine [Canadian Science Centre for Human and Animal Health]