A comprehensive insight on Nipah virus, its characteristics, historical outbreak including recent Kerala outbreak and prevention and control measures.

1. NipahVirus: An Introduction
From Fear toTragedy
Delivered by
Dr. DeepthiVijay
Assistant Professor,VPH, KVASU

2. About theVirus
• Family- Paramyxoviridae
• Order- Mononegavirales
• Paramyxoviruses: -ve sense, ssRNA viruses
• Cause high human & animal health burden
(e.g., measles, distemper, mumps, parainfluenza,
Newcastle disease, respiratory syncytial virus
and metapneumoviruses etc.)
• Traditionally these viruses have:
– Narrow host range
– Low mortality rate (Virtue et al., 2009)
 Vaccination- reduced impact of diseases
(measles & mumps) (Holzmann et al., 2016)
 Eradicated Rinderpest : Imp. livestock disease
(Mariner et al., 2012)

3. Emergence of Henipavirus
• Various ecological & anthropogenic factors
 Spill over of diverse & novel paramyxoviruses
that circulate in wildlife reservoirs  to other
terrestrial mammals, including humans
• Emergence of two highly pathogenic viruses
– Hendra virus
– Nipah virus
• In 2002, International Committee forVirus
Taxonomy (ICTV) approved the est. of genus
Henipavirus due to distict genetic and
molecular features (Eaton et al., 2006)

4. Hendra virus (HeV)
• 01st zoonotic henipavirus to be identified
• In 1994, outbreak of hendra virus was
reported in horses in Australia
• 02 people in close contact with these horses
developed febrile illnesses (01 died)
• Reservoir: Australian flying foxes
Virus Spillover
• Flying foxes-to-horses
(respiratory & neurological disease)
• >50 HeV outbreaks in horses (Middleton, 2014)
• Horse-to-human via. close contact
• Human- 07 cases/ 04 deaths (Mahalingam et al., 2012)
• Human-to-human: Not observed
(Playford et al., 2008)

5. A succinct of Nipah outbreaks
• 1st outbreak-1998, Malaysia- respiratory and neurological disease in
pigs and encephalitis in people (Chua, 2012)
– Importation of infected pigs into Singapore from Malaysia  a cluster
of infections in abattoir workers in Singapore (Chew et al., 2000)
• 03 years later (2001), geographically distinct NiV strain
independently emerged in Bangladesh as well as in India
• 2014- Philippines- transmission horse-to-human (ching et al., 2015)

6. Nipah virus characteristics
• Morphology: pleomorphic (spherical to filamentous) ranging in size from
40-1,900 nm (Wang et al., 2013)
• 1st isolate was recovered from clinical material from a fatal human case
from Sungai Nipah (Malaysia) in 1998 (Chua et al., 2000)
Two strains:
• Malaysian strain (NiV-MY)
• Bangladesh strain (NiV-BD)
# Philippines outbreak (2014) was most likely caused by NiV-MY

7. Major pathways of Nipah virus infection
Malaysia (blue arrows); Bangladesh (red arrows), Philippines (green arrows)
(Clayton, 2017)

8. Bat as Reservoir
As the flying fox (fruit bat) habitat is
destroyed by human activity
- bats get stressed & hungry
- immune system gets weaker
- virus load goes up
Virus spills out in their urine & saliva
• Broader distribution of NiV in
Pteropid fruit bats across their range
(Reynes et al., 2005)
• No clinical signs in bats (Wang et al., 2013)
• Viruses related to HeV & NiV also
circulate in non-pteropid fruit bats
across the globe (Clayton et al., 2013)

9. Pteropus sp. distribution Map
(Ang et al., 2018)

10. VirusTransmission
02 mechanisms of virus spillover from flying foxes:
– Transmission via intermediate animal host - Malaysian outbreak
– Bat-to-human transmission & human-to-human transmission
(Bangladesh and India)
• Two most common risk factors for human NiV infection was
– drinking contaminated date palm sap/ fermented product
– contact with a patient with nipah encephalitis (Hedge et al., 2016)
Bangladesh
& India
Malaysia

11. Epidemiology in Animals
• Host range- Broad species tropism
• In addition to multiple species of bats, NiV naturally infected pigs,
horses, dogs, cats, goat and humans in the outbreak areas of
Malaysia but all were effectively dead-end hosts (Wang et al., 2013)
• This wide range of species tropism is in part due to the fact that
NiV uses ephrinB2/B3 molecules as their entry receptors, which are
highly conserved amongst all mammals (Negrete et al., 2006)
(Dutta et al., 2016)

12. Lessons from the past…
Malaysian Outbreak- 1998

13. • Studies using satellite telemetry  Highly mobile Malaysian flying
foxes can travel hundreds Kms between roosting sites in a year
• Home ranges- beyond Malaysia to include Indonesia & Thailand
(Epstein Jonathan et al., 2009)

14. Epidemiology of the Outbreak
• In 1998, Malaysian outbreak
– Intensively farmed pigs, sickened by fruit bats
– Slash-and-burn deforestation
• 1st isolated from Sungai Nipah NewVillage (Field et al., 2001)
Intensive
animal
agriculture
Long-distance
animal
transport
Habitat
destruction
(WHO/FAO/OIE report)
Interplay of multiple ecological risk factors

15. • In 1997-98: >10mn acres forest burnt in Borneo & Sumatra
• Exacerbated by El‟Nino  drought, haze and habitat loss
• Unprecedented encroachment on orchards resulted in porcine
exposure to flying fox saliva or urine (Chua et al., 2002)
• Rapid transportation of infected pigs- 05 states of Malaysia &
Singapore resulted in 265 human cases (105 deaths) (CDC, 1999)
• Culling of > 1 mn pigs contained the catastrophe (Lam, 2003)
Mass exodus of “flying fox” for searching for food

16. Animal Husbandry- conducive factors
• Index farm triggering the large outbreak was one of the largest
hog operations in the country with ~30,000 animals (Ludwig et al., 2003)
• Despite being an Islamic country  annual pig production had risen
significantly in the decade prior to outbreak-
~ 3million head to ~ 5million (FAO 2007)
• Other factors:
– population size & stocking density of the pigs
(Nipah presumably a density-dependent pathogen- respiratory route)
– continual influx of young, immunologically naive pigs (Olival & Daszak, 2005)

17. Outbreak analysis
• 03 outbreaks in Malaysia- Sept.,1998 to Jan.1999 (Tan et al., 1999)
• Initially, the cases were considered as Japanese B Encephalitis (JE) -
which previously caused porcine-associated outbreaks in Malaysia &
JE nucleic acids were detected in patients‟ sera (Chua, 2003)
• Initial measures  fogging to kill mosquitoes & JE immunization

18. JE vs. Nipah
Confusing transmission cycle and clinical symptoms
Nipah

19. JE vs. Nipah during Malaysian outbreak (Chua et al., 1999)
– Most patients were adult males rather than children
– High proportion of victims had direct physical contact with pigs,
unlike a mosquito-borne disease
– Clustering of symptomatic cases among members of the same
household was as high as 33% (Tan et al., 1999) suggesting an attack rate
higher than that of JE virus, which is symptomatic in only 1: 300
infected (Thongcharoen, 1989)
– Many patients had previously been immunized against JE
– Infected pigs developed a severe barking cough and many dying from
the disease (not a feature of JE) (Looi & Chua, 2007)
# In pigs the disease is also known as „„barking pig syndrome‟‟ & „„one-mile cough‟‟

20. Detection ofVirus and its reservoir
• In March 1999, virologists from University of Malaya isolated a virus
that belonged to Paramyxoviridae and sequencing at CDC proved
that it is about 20% different from the Hendra virus (Chua et al., 2000)
• The fruit bats of Pteropid sp. were identified as the natural reservoir
hosts of the virus and its transmission to pig (Yob et al., 2001)
• Fruits eaten by bats may have been dropped into pig sties and
subsequently infected the pigs that consumed contaminated fruit

21. Measures to contain the outbreak
• Health education to pig farm workers on
(Ang et al., 2018)
• Personal protection
• Handwashing after handling of animals
• Biosecurity and disinfection of farms
• Disease control in Pigs (Lam & Chua, 2002)
– Pig-culling operations
– Phase I - culling in areas where outbreak
cases had occurred
• > 1 million pigs (45 % of all pigs in Malaysia)
were culled
– Phase II - surveillance in all pig farms
throughout the country
• Farms at which ⩾3 samples had positive
results for NiV were considered to be
positive farms
• All pigs at the affected farms and at farms
within a 500-m radius were culled
• This process was carried out for 3 months

22. Outbreak in Singapore
Feb.1999- Malaysian outbreak spread to Singapore through imported pigs
• 04 cases of encephalitis were noted among abattoir workers
• NiV was identified by RT-PCR in CSF/ tissue of patient & the sequence
was found to be identical to sequences from Malaysian cases and pigs
(Chew et al., 2000)

23. Control Measures
• > 500 abattoir workers were screened
in following week and those with
symptoms of respiratory or
neurological disease were admitted to
hospital
• The 02 abattoirs in Singapore closed
down for investigations and disinfection
• The outbreak ended with the ban on
importation of live pigs and its
products from Malaysia

24. Outbreak in Bangladesh and India
Bangladesh
• 2001- outbreak of encephalitis in Meherpur
• Not investigated until 2003- when another cluster of febrile illnesses
with neurologic features with 08 deaths in adjoining villages in
Naogaon district (≈150 km from Meherpur)
India
• Feb. 2001- outbreak of febrile illness associated with altered
sensorium in Siliguri,W.B., India
• Lab. investigations did not identify any known infectious agent
Retrospective investigation of Bangladesh oubreak was conducted
in March 2003  samples were sent to CDC  +ve NiV
(Hsu et al., 2004)
• Siliguri  near to Bangladesh  retrospectively analyzed  +ve NiV
(Chadha et al., 2006)
• Since 2001-NiV outbreaks have continued to occur in Bangladesh
and India on almost annual basis

25. Epidemiology of the outbreak…
• Unlike in Malaysia and Singapore, pigs were not involved as an
intermediary host
• Bangladesh is a predominantly Muslim country, with no pig farming,
while in W.B, India, though there are pig farms, it is not on the same
scale as in Malaysia

26. • In Bengali culture- sap harvested from date palm
tree is commonly used for fresh consumption/
fermented into alcoholic drinks (toddy, tari or palm
wine)
• Pteropus spp. bats frequently feed on the shaved
bark and often contaminate the sap with saliva,
urine, and excreta
• Consumption of fresh date palm sap as the primary
route of bat-to-human transmission
• Other risk factors
– Climbing trees (probably contaminated with
infected date palm sap)
– Contact with sick animals
– Fruit bats commonly drop partially eaten saliva
laden fruit which are then eaten by domestic
animals foraging for such food (Islam et al., 2016)
Epidemiology of the outbreak…

27. Person-to-person transmission
Nipah virus infection of health care workers during the outbreak
characterized by nosocomial transmission
• Exposure to respiratory secretions of patient during procedures
such as intubation, and failure to wear PPE when in contact with
infected patients (Chadha et al., 2006)
• Specific cultural practices
– Transmission between infected patients & their caregivers
• Societal norms - care is provided by patients‟ family members even
in hospital settings, where trained health-care professionals adopt a
“hands off” approach to patient management (Hadley et al., 2007)
– Number of cases of human infection were attributed to corpse-
to-human transmission (embracing body of a loved one soon
after their death, or ritual preparation of a corpse for burial)
(Sazzad et al., 2013)

28. Outbreak in Philippines
• 2014- Philippines National Epidemiology Center
received a report of human deaths in 02 villages
on Mindanao
• Outbreak investigation revealed additional
human deaths with concurrent neurologic
disease and sudden deaths in several horses
• Neutralizing Abs against NiV in horses and IgM
against NiV in 3 patients
Virus transmission to humans
• Direct exposure to infected horses
• Contact with contaminated body fluids
during slaughtering of sick horses
• Consumption of undercooked meat of
infected horses
• Overall fatality- 53% (Ching et al., 2016)

29. Clinical Signs of Nipah in Human
• I. P: 4 days to 2 months (> 90% at 2 weeks or less) (Goh et al., 2000)
• Patients presented with
• Fever
• Headache and dizziness
• Vomiting
Which developed into a picture of
Severe encephalitis
• Reduced level of consciousness
• Prominent signs of brain-stem dysfunction
• Neurological involvement- diverse and multifocal
• Cerebellar signs were common
A unique and interesting feature - development of relapse and late
onset encephalitis (some occurred months/ years after acute illness)
Respiratory Involvement
• Malaysian outbreak- respiratory involvement- 14-29% of cases
• Bangladesh and Indian outbreak- higher rates of respiratory involvement
• ½ to 2/3 cases- develop acute respiratory distress syndrome

30. Signs of brain-stem dysfunction
MRI of multiple small white matter lesions
(A) Multiple punctate white matter lesions (arrowheads)
(B)The largest lesion is more prominent on corresponding
diffusion-weighted image (DWI)
Virus Nipah in CSF of infected
patient

31. Parameter Malaysia-Singapore Bangladesh-India
Age and occupation Mainly adult pig farm workers Adults, children & healthcare
workers
Spread •Bats-to-pigs,
•Pigs-to-human
•Occasionally:
Human-to-human
• Bats-to-human by contaminated
date palm juice and fruits
• Possibility of bats-to-domestic
animals-to-humans
• Human-to-human spread
Respiratory
involvement
•14-29% Malaysian cases
•2 /11 patients in Singapore
had pneumonia without
encephalitis
Cough (62%), respiratory difficulty
(69%); chest radiographs with
acute respiratory distress
syndrome
Encephalitis Segmental myoclonus seen in
32-54%
Segmental myoclonus not
reported
Relapsed and late
onset encephalitis
About 5-10% Delayed onset neurological
abnormalities in 4/22 patients
Persistent
neurological deficits
About 20% About 30%
Mortality 32-41% 70 %

32. Treatment measures
• Treatment - largely supportive consisting of
– Anticonvulsants
– Treatment of secondary infection
– Mechanical ventilation and rehabilitation
• Malaysia outbreak- empiric treatment with ribavirin (broad
spectrum and ability to cross the blood-brain barrier)
• In an open-label trial of ribavirin- a reduction in mortality observed
(54% in control vs 32% treatment) (Chong et al., 2001)
• Malaysia outbreak- Mortality rate: ~ 40%
• Bangladesh and India- Mortality rate ~ 70%
 Greater involvement of respiratory tract in Bangladesh & India outbreak
 Differences in pathogenicity between 02 viral strains
 Less advanced healthcare facilities (Intensive Care Units) in Bangladesh

33. Details of current outbreak in Kerala

34.  Nurse Lini Puthussery: contracted infection while treating the
first 03 victims of infection
The Great Martyr - Lini Puthussery
To prevent the spread of outbreak  her body was cremated in electric
crematorium immediately without handing it over to her family

35. A letter to husband…..

36. A panic among public !!!!!
Calicut Medical College

37. How doctors spotted Nipah ????
G.S. Mudur (May 25, 2018)
Anoop Kumar AS, Chief,
Critical care unit BMH
17th May @ Baby Memorial Hospital, Kozhikode
• Muhammed Salih (26 yrs) – admitted with
fever, high B.P and acute confusion
– His B.P increased and was unable to
breathe- doctors had to intubate him
# Doctors suspected either a case of poisoning
or a deadly infectious virus
# Patients with severe encephalitis are at risk of
falling B.P but here the pressure seemed to
be rising- Anoop Kumar, AS
• Salih's aunt Mariyam also had high fever and
was admitted to BMH
• Within 24 hours, Salih suffered a fatal cardiac
arrest due to inflammation of his heart and
Mariyam died the next day
• Just 12 days earlier, Salih's 23-year-old brother
had died with similar symptoms

38. “Nothing seemed to work.There was another patient in the family (a 50-
year-old woman) with similar symptoms, and the patient's brother had died
similarly a few days earlier”
The brother, believed to have been the first patient (Index case),
had fallen sick on 2nd May and died on 5th May
"Immediately, we got on alert that we are dealing with some abnormal viral
encephalitis; it is not the normal one," - Kumar AS
The samples were sent to
Manipal virology centre

39. Arunkumar G.
Head Manipal Center forVirus
Research
• @ Manipal virology centre – Dr Arunkumar sensed
from case descriptions that the infection was
unlikely to be one of commonly cause encephalitis
• “Japanese encephalitis doesn't occur within a
family, either - you see about one case in a village
during an outbreak"
• "The rapid deterioration and the cluster from a family
were warning signs of nipah“
• Scientists at Manipal screened Salih's samples for 35
common agents of encephalitis and found Nipah +ve
• The findings were confirmed by independent analysis
by National Institute ofVirology, Pune, which arrived
on May 20.
• NIV is now undertaking full genome sequencing and
isolating the virus in its state-of-the-art Biosafety
Level 4 lab

40. Kudos to Kerala multi-pronged approach
containing the outbreak so quickly….
• Within 48 hours- state health officials
and central government had diagnosed
the virus as Nipah
• The state's emergency health system
kicked in and the outbreak, for now,
appears to be controlled
– Although, the virus has avg. 14-day
incubation period, so it'll be a month
before scientists can say the outbreak has
been fully controlled)
• The 11 fatalities have been linked to
Salih's family
• "Everybody is linked, there is no
unlinked case, that is very satisfying” –
Arunkumar
• 2001 outbreak of Siliguri,W.B. took
05 years to identify the virus

41. SourceTracking….
• The first three cases were from a single family in Perambra
– Muhammad Salih, 26
– his brother Muhammad Sabith, 23
– and his paternal aunt Mariyam, 50
• The brothers were cleaning a well that was home to bats

42. • The animal health department is collecting samples of rats,
bats and other species to find the Nipah reservoir host
• Samples are being analyzed by the NIHSAD, Bhopal
Is there any role of Intermediate animal host
or
human reservoirs (migratory population) ????

43. • Recent Nipah outbreak can cause a knee-jerk reaction of calls
for bat culling….
• Studies warn that instead of reducing the outbreak of such zoonoses,
it could cause even more damage- mainly ecological damage
• About a quarter of >1,300 bat species seen worldwide feed on fruit
and nectar and are crucial pollinators, helping maintain genetic
diversity in agricultural systems
• They are also important seed dispersers; other bat species help bring
rodent and insect numbers under control

44. INDIA- a hotspot for zoonoses
• Fast-growing human
population
• Increasing animal-human
interactions
• Changing environmental
conditions
• Inadequate sanitation and
regulation
• Socioeconomic and
anthropogenic factors

45. Corollary: According to (severely underreported) official data:
In 2017:- J.E ("villain" in Gorakhpur) killed 254 while Non-JE AES killed 1097
(Actual could be 50-100 times more)
Of these, how many could have been #Nipah deaths????

46. Biosafety Issues during Nipah outbreak
• Field and Farm workers should use PPE (masks,
goggles, gloves, gowns, and boots) together with
hand-washing and disinfection (FAO, 2002)
• NiV fulfils criteria of a potential
bioterrorism agent
– high virulence
– significant morbidity and mortality
– resultant fear and panic
– tremendous economic losses caused (Lam, 2003)
• NiV Listed as a Category C agent by CDC
• Virus handling must be done in BSL-4 facilities

47. Samples to be collected
Standard Triple Packaging
• Blood, throat swabs, urine and Cerebrospinal fluid (CSF) samples
can be collected
• At least 5 mL of whole blood should be collected in a plain tube
• Throat swabs should be collected and transported inViral
Transport Medium (VTM) in a sterile, screwcapped polypropylene
tube
• About 10-20 ml of urine samples should be collected in to a
sterile, screw-capped, leak-proof and sturdy container.
• About 2-3 ml of CSF specimens should be collected by lumbar
puncture into a sterile tube, and The CSF specimen should not be
frozen
• Samples for PCR testing should preferably be sent in dry ice, or
at -20°C, using hard-frozen gel packs
• Tissue biopsy specimens, if available, should be transported in a
screw-capped, leak-proof container with a small volume ofViral
Transport Medium, preferably in dry ice or at -20°C.
Guidelines,NIV, May-2018

48. Collection and transportation of samples
Sample vials & other accessories Suitable sample containers, gel packs & coolant box
Arrange the sample vials with proper labeling Place vials in leak-proof secondary container
Arrange the packed container in outer shipment container with case report form & labeled address

49. Diagnosis of NiV
• Infections by NiV in humans and animals are confirmed by virus isolation,
nucleic acid amplification tests and serologic tests
• For isolation and propagation: BSL-4 facilities are needed
• The OIE reference lab. for Henipaviruses in Asia-Pacific region is located at
Australian Animal Health Laboratory, Geelong
• Bangladesh NiV outbreaks are handled by ICDDRB and IEDCR in
collaboration with CDC, USA
• In India : BSL 4 lab at National Institute ofVirology (ICMR), Pune
– High Security Animal Disease Laboratory, Bhopal with BSL3 + facility caters the need for
exotic animal disease diagnosis
• Infections by NiV in humans and animals can be confirmed by serological
tests (the most commonly used serologic assays are ELISAs)
• NiV infection can be detected by molecular diagnostic tests like RT-PCR,
Real time RT-PCR (Taqman) and Duplex nested RT-PCR which can be
confirmed by sequencing of amplified products

50. Prevention and Control
• Early detection of outbreak and installing preventive measures as soon
as possible is very imp. as treatment options are limited
• Medical and veterinary professionals along with ecologists should
increase the awareness of the disease particularly hosts and mode of
transmission of the virus- “one-health” approach
Preventive strategies
@ farm animal level
 Interventions to prevent farm animals from acquiring NiV via direct
contact of bats or eating fruit contaminated by bats
 Farms should be designed to reduce over-crowding to avoid rapid
spread of disease and should not be near fruit trees which attract bats
@ food contamination
 Consumption of contaminated fruits/ sap should be avoided
 Physical barriers to prevent bats from accessing & contaminating sap
@ human-human-spread
 Avoid close contacts with infected individuals
 Proper use of PPE
 Proper Hand hygiene
 Consult to physician in suspected cases

51. Public Awareness

53. Why collaboration??
The Six Blind Men and the Elephant

54. In nutshell: What model we need ???
Multiple expertise needed !
Infectious
diseases
Surveillance
Units
Inter-sectoral
Public Health
Units
Laboratory
&
Diagnostic
Services
MolecularTools
GIS &
Bio-informatics
Epidemiological
Investigation
teams
Information
sharing
management &
rapid action
International
Collaborations

55. “A ounce of prevention is
wroth a pound of cure”
–Benjamin Franklin
Contact Mail:
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