Challenges of New and Challenges of New and emerging infectionsemerging infections

Prof.N.K.Ganguly,Director General, ICMR2nd Asian Science and Technology Forum,

Tokyo, Sept. 8th 2006

MRCIRMS

NICED

TRC

EVRC RMRC

RMRCCRME

NIE

VCRC

CJIL

RMRI

NARINIV

DMRCRMRC

RMRC

ICMR Institutes Researching in Communicable Diseases

Siliguri encephalitis, 2001Siliguri encephalitis, 2001an explosive outbreakan explosive outbreak

Cases, mainly adults, of fever with altered sensorium in differCases, mainly adults, of fever with altered sensorium in different ent hospitals at Siliguri W. Bengal between 31hospitals at Siliguri W. Bengal between 31stst January through February January through February 2001.2001.

45/66 (68%) patients health workers or visitors; CFR = 73.8%45/66 (68%) patients health workers or visitors; CFR = 73.8%NIV-ICMR

0

2

4

6

8

10

12

31 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23Date of Onset

No.

of c

ases

Associted with Hospital Association not establishedNo. of Deaths

JanuaryFebruary

Nipah BanSILI-N2SILI-N1

SILI-N4SILI-N5

SILI-N3Nipah3 MalaysiaNipah6 Malaysia

Nipah1 MalaysiaNipah2 Malaysia33

3099

4562

88

0.005 Nipah2 Malaysia

NIPAH virus etiology

Nipah1 MalaysiaNipah3 Malaysia

Nipah1 BanNipah2 Ban

SILI-M177

100

0.005

• N gene amplified from 5/6 urine samples. (99.95%, & 97.11% homology) • M gene amplified in 3/6 samples (98.91% and 94.37% homology)

Nipah virus etiology confirmedChadha et al., 2005 EID

• Negative for antibodies to JE, WN, Den, Measles

CDC collaboration• 9/17 cases +ve IgM abs, 10/17 +ve for IgG abs for Nipah virus

Clinical featuresClinical features

Acute fever, mild to high grade (100Acute fever, mild to high grade (100--104 F)104 F)

Pediatric age group below 15 yrsPediatric age group below 15 yrs

Altered sensorium, Altered sensorium, camatosecamatose condition leading to death in condition leading to death in 11--2 days2 days

High mortality (CFR ~ 60%)High mortality (CFR ~ 60%)

No neck rigidity, No neck rigidity, KernigKernig’’ss sign absentsign absent

Malnourished children belonging to low socioMalnourished children belonging to low socio--economic economic class and poor hygieneclass and poor hygiene

Saharanpur outbreak 2002-04

A total of 40 cases with 25 deaths A total of 40 cases with 25 deaths (CFR (CFR 63%) 63%) reported from 12reported from 12--1010--02 to 1202 to 12--1212--0202

A total of 34 cases with A total of 34 cases with CFRCFR 80% 80% reported during Septreported during Sept--Oct 2003.Oct 2003.

During 2004, a total of 158 cases with 113 During 2004, a total of 158 cases with 113 deaths deaths (CFR 72%)(CFR 72%) reported between reported between OctoberOctober--December.December.

A few cases positive for viruses like A few cases positive for viruses like adenaden, , enteroentero etc.etc.

Saharanpur outbreak 2002-4

Negative for all viruses in culture and PCR

Diagnosis--Inconclusive

EM: Adeno-like particle from CSF

SARS Outbreak, 2003

MolecularPolymerase Chain Reaction (PCR)

Cell cultureElectron Microscopy

1 2 3 4 5 6 7 8 9 10 11

Global SARS Outbreak, 2003

June to September 2003, 319 cases of encephalitis with 174 deaths (CFR 54.5%) were reported from 11 districts of A. P.

Children below 14 years age, predominantly belonging to low socio-economic group, rural areas

Total chaos and panic as onset of disease was abrupt, 85% of deaths within 24 hours of hospitalization

All samples negative for known causes of encephalitis

Chandipura encephalitis outbreak, AP 2003

virus isolations:10strains in RD, Vero, MDCK cell lines and infant mice

Laboratory Studies

FA: CHP viral antigen in brain cells confirmed

PCR: CHP G gene primers amplified RNA from isolates and clinical samples. 98% homology with 1965 isolate.

Electron microscopy: Bullet shaped particles and helical nucleocapsid - typical Rhabdoviruse.

Identifications: Chandipura virus by CF and NT and PCR

Isolations, PCR and serology suggested involvement of CHP virus in these encephalitis cases

Subclinical infection recorded

Chandipura virus was found responsible for the outbreak in AP and adjoining districts of Maharashtra

Rao et al 2004; Lancet

An outbreak among tribals from Chotaudepur and Kawat Talukas in Vadodara dist and 1 taluka in Panchmahal district.

26 cases with 17 deaths (CFR 65%) between 9-6-04 to12-7-04.

80% of the deaths within 24 hours of hospitalization.

Chandipura encephalitis in Gujarat, 2004

CHP sequences: closely related to prototype strain (1965) and Andhra Pradesh (2003) isolates.

It was concluded that the etiological agent was Chandipura virus

Vadodara2Vadodara84

Vadodara125Vadodara103

Vadodara14Vadodara109

AP18RAP31M

Vadodara6Vadodara115AP27M

Vadodara112AP09R

AP60VMAH6514V96

5269

4943

76

4935

5033

55

31

0.01

Clinical samples –ve for JE, West Nile, Dengue and Paramyxovirus

Virus isolations : one in RD, PS Cell lines and in suckling mice.

PCR: CHP RNA in 9/20 (45%) acute serum samples.

ELISA: 2 patients +ve for CHP anti IgM abs.

Seroconversion: 1 case –ve for IgM in first sample turned +ve in second sample

Laboratory investigations

Chadha et al 2005; American J Trop Med Hyg

Chandipura encephalitis in A P, Chandipura encephalitis in A P, 20052005

PeriodPeriod 44thth May May –– 3131stst

AugustAugustTotal recorded Total recorded deaths/ casesdeaths/ cases

32/54 32/54 (59.3%)(59.3%)

Serum samples Serum samples from probable from probable cases cases ##

35 (65%)35 (65%)

Probable cases Probable cases deaths/casesdeaths/cases

21/35 (60%)21/35 (60%)

# encephalitis cases in age group < 15 yrs, fever of less than 5days at admission, and CNS involvement

Etiology was confirmed as Chandipura virus by isolation, seroconversion and PCR

Warangal, Karimnagar, Khamam and Adilabad districts

In India, Isolated from sandflies in1970 in Aurangabad dist. of Maharashtra and 3 isolations from AP

Isolations from hedge hog in Nigeria, and sand flies in Senegal, West Africa,1992.

In sri Lanka, 2/115 monkeys positive for abs

Sandflies are probable vector & extra human reservoir may be present

CHP: Vector distribution, ecology

Acute Acute HaemorrhagicHaemorrhagic fever in Mumbai, 2005fever in Mumbai, 2005

KalyanKalyan //DombivaliDombivali area, August, area, August, 2005. 2005. AdolescentAdolescent-- adults with short history adults with short history of fever, bloody vomit and death of fever, bloody vomit and death within short duration within short duration Long exposure to stagnant water Long exposure to stagnant water during floods.during floods.Family contacts spared.Family contacts spared.Post mortem in 3 cases Post mortem in 3 cases -- grossly grossly hmghmg lungs, kidneys and necrotic liver. lungs, kidneys and necrotic liver. Sera and organs collectedSera and organs collected

Laboratory investigationsLaboratory investigations

Samples inoculated in suckling mice and cell culture Samples inoculated in suckling mice and cell culture -- no no isolations.isolations.

DengueDengue1of 51 sera samples positive 1of 51 sera samples positive All sera from 12 dead cases were All sera from 12 dead cases were negaivenegaive ..

LeptospiraLeptospira3/12 cases positive for 3/12 cases positive for LeptospiraLeptospira by MAT; PCR for by MAT; PCR for LeptospiraLeptospira negative. negative.

HantaanHantaanAll sera from 12 dead cases were negative.All sera from 12 dead cases were negative.PCR for Hantaan using PM organs were negative.PCR for Hantaan using PM organs were negative.Sera from contacts of dead cases +Sera from contacts of dead cases +veve for IgM abs for for IgM abs for Hantaan by commercial kits. Hantaan by commercial kits.

No viral etiology; Leptospirosis??

• Avian Influenza outbreak recorded in Maharashtra

• Poultry census, culling and tracing missing poultry

• Total destruction of backyard poultry

• Following outbreak in poultry >500 samples from handlers, cullers, contacts collectedOutbreak of Avian Influenza in Maharashtra, 2006

Rapid Molecular Tools in Virus Rapid Molecular Tools in Virus Identification: Avian FluIdentification: Avian Flu

NASBA results detecting very low amount of H5N1 RNA

Virus detection by Real time PCR

Real Time PCR System

State-of-artNASBA Platform

H5N1 gene of avian influenza virus detected by RT-PCR

Samples from suspected cases were processed for H5 and N1 PCR; all negative

All samples processed by Real-Time PCR/ NASBA –negative except 1 environmental sample that gave positive signal for N1 gene in NASBA

Samples were inoculated in eggs/ MDCK cell lines-no isolations

No sample met the criteria-hence all negative

Avian Influenza in Humans, 2006

Sudden onset of fever and crippling migrating arthralgias,which may last for a few days to months.

Papular or maculopapular skin rash may occur, typically on the trunk and limbs.

Normally, self resolving disease

Complications include-

Mild hemorrhagic manifestations

Meningo-encephalitis, Encephalitis

Maternal-foetal transmission

The relation between Chik and death is still under investigation.

Chikungunya outbreak, 2005-06

Chikungunya virus activity in India

60s to 80s 2005-06

Gujarat

Rajasthan

Madhya Pradesh

Maharashtra

Andhra Pradesh

Karnataka

TamilNaduKerala

Orissa

Chikungunya Laboratory Diagnosis

Virus isolationsImmunofluorescence Assay ( IFA )

RT-PCR for genomic detectionMAC ELISA

1.75 kb

2.0 kb

2.1 kb 1.3

kb

365 bp

Sporadic activity in India (2000-2004)

PlacePlace MACMAC--ELISAELISA--PositivePositive

HI PositiveHI Positive PCR PCR positipositiveve

KolhapurKolhapur 3/1203/120 2/112/11 ++

NandedNanded 2/112/11 1/161/16 --

GadchiroliGadchiroli 2/332/33 4/174/17 ++

PCMCPCMC 3/823/82 1/101/10 --

SuratSurat 1/1161/116 0/280/28 --

KeralaKerala 7/947/94 NDND NDND

HydrabadHydrabad 3/473/47 NDND NDND

Total no. of samples tested by MAC-ELISA for CHIK Infection [n=865]

ChikChik-- vector in Indiavector in India

• Ae aegypti is the main vector

• Virus is transmitted by bite of female mosquitoes

• Primarily a daytime feeder• Lives around human

habitation• Breeds in artificial

containers

0.02

YawatCentral Africa

(1982-96)

Indian Ocean (2005-06)

South/East Africa(1952-76)

Asian(1963-83)

West Africa(1964-83)

Congo(2000)

UG-82

CHIK partial E1, (1044 nt)

All current isolates belong to African genotype in contrast to earlier Asian isolates

PhylogeneticPhylogenetic relationships among CHIK isolates based relationships among CHIK isolates based on E1 sequenceson E1 sequences

CONGOAF192906AF192907

YawatAY549583

CENTRAL AFRICA

RajasthanVelloreAPParbhaniBangaloreAM258990AM258991MauritiusReunionAM258993AM258992AM258994

SOUTH/EAST AFRICAASIAWEST AFRICA

100

100

100

98

99

89

29

25

79

8468

44

100

47

43

49

55

44

CONGOAF192906AF192907

YawatAY549583

CENTRAL AFRICA

RajasthanVelloreAPParbhaniBangaloreAM258990AM258991MauritiusReunionAM258993AM258992AM258994

SOUTH/EAST AFRICAASIAWEST AFRICA

100

100

100

98

99

89

29

25

79

8468

44

100

47

43

49

55

44

AP-1AP-2KAR-1KAR-2MAH-1MAH-2

AF345888ROSS (Africa)S27-1 (Africa)S27-2 (Africa)

BARSI-73CHENNAI-64NAGPUR-65-1CHENNAI-71NAGPUR-65-2

KOLKUTTA-63-1KOLKUTTA-63-2SENEGAL-83

4991

92

6640

67

91

0.02

Emerging Inf Diseases, Oct 2006

Reunion

AM258994

AM258993

AM258992

AM258990

AM258991

Bangalore

Parbhani

Rajasthan

Vellore

AP

Yawat

ROSS

S27-2

S27

MOS-Senegal-83

ONY-UG96

100100

100

100

537581

86

100

100

6480

92

0.02

Reunion

AM258994

AM258993

AM258992

AM258990

AM258991

Bangalore

Parbhani

Rajasthan

Vellore

AP

Yawat

ROSS

S27-2

S27

MOS-Senegal-83

ONY-UG96

100100

100

100

537581

86

100

100

6480

92

0.02

PhylogeneticPhylogenetic relationships relationships among CHIK isolates among CHIK isolates

based on based on whole genome whole genome sequencessequences

Whole Genome to study microevolution and pathway analysis?

Diagnostic Electron Microscopy

Virus morphodiagnosis

Immunoelectron microscopy

Cryoelectron microscopy

Global networking

EQA accredition & resource sharing

Telemicroscopy links

Poxviruses

UAcUAc AMdateAMdate PTAPTA

Lesion from farmer’s hand direct TEM imaging of vesicular fluid

Parapoxirus

Gastrointestinal virusesSporadic outbreaks, cases

Adeno Rota

Astro Calici

Fecal specimens, direct imaging after negstain

Sapporo and Noroviruses

Rapid ultrathin sectioning K4M

CPE+ cultures BHK21 cells

TEM characterization of some unclassified viruses from India

A Bunyavirus

Chittoor virus

0.1

SAB1VP1KEKZK00018

KAGBG00024KABLK00002

KAGBG00022KAKPP00004

UPMRT01029UPMRT02021

UPMRT01041UPMRT01038

UPMTR02012UPMRT02017

UPMRT02056UPPIL02044

UPMZN02103UPMZN02076

UPMZN02055UPMZN02044UPMZN02084

UPMRT02016UPMRT02002

UPMZN02035UPMRT01037

UPMRT01108KABLY03006

UPMRT02038

Introduction of wild poliovirus type 1 from UP to Karnataka, 2003

Enterovirus Research Centre, Mumbai ICMR, New Delhi

Division of Epidemiology and Communicable Diseases

Polio Eradication Initiative June 2005

Transmission pathways of wild polio virus 2004-2005

Case or outbreak following importation

Endemic countries

Wild virus type 1Wild virus type 3

Re-established transmission countries2 0 0 2

11

162

45

0 0 0 114

5

0

20

40

60

80

100

120

140

160

180

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05

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05

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05

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05

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r

05

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YEMINO

AngolaIndonesia

Wild Polio1 Importation in Angola

Importation of wild poliovirus 1 in Indonesia

18 Jan 06299 cases

M

F

G

14

16 G ward

M warda

International collaboration in Emerging International collaboration in Emerging Infectious Diseases and Disease Infectious Diseases and Disease

SurveillanceSurveillance

Sharing of informationSharing of informationBiological Material for R&DBiological Material for R&DCollaborative researchCollaborative researchSite preparation for clinical trialsSite preparation for clinical trialsInvestigation of outbreaksInvestigation of outbreaksInfrastructure strengthening esp. for labs of Infrastructure strengthening esp. for labs of appropriate appropriate biosafetybiosafety levelslevelsTraining for managing outbreaks and Training for managing outbreaks and biosecuritybiosecuritylevelslevelsWorkshopsWorkshops

RecommendationsRecommendationsIncreasing research capacities and capabilities of Increasing research capacities and capabilities of developing countries to deal with emerging infections.developing countries to deal with emerging infections.Human networking as important as technology based Human networking as important as technology based networking to address emerging infections.networking to address emerging infections.Need to develop networking between research Need to develop networking between research organizations and institutions involved in emerging organizations and institutions involved in emerging infectionsinfectionsA regional network of networks would help in sharing of A regional network of networks would help in sharing of informationinformationICMR to be nodal agency for emerging and reICMR to be nodal agency for emerging and re--emerging emerging infections in Indiainfections in India