UNDERSTANDING THE COMPLEXITY OF GENES AND DISEASES IN INDONESIAN ARCHIPELAGO

Herawati SudoyoEijkman Institute for Molecular Biology

Jakarta, IndonesiaHealth systems complexity: Bridging Physiome with Health Care through

Computational Modelling Symposium, NITH-NTU, Singapore, 26 January 2015

GENNEKA TUNGGAL IKAUnity in diversity

Indonesia and Infectious Diseases – a Great Challenge to Mitigate Biorisk

§ Problems with emerging and reemerging infectious diseases

§ Most caused mainly by environmental, ecological or demographic factors spread by travel and trade – Indonesia is a maritime country with 17.504 islands, 700 languages, 33 provinces, 230 million population

Problems with people movementRecognize the need to develop, strengthen and maintained the capacity to detect, report and respond to public health events

WHAT ARE WE FACING?

§Indonesia – A very diverse populations –vast genome diversity – disease management complex §Indonesia - A rapidly developing country with serious challenges in infectious (emerging and re-emerging) and zoonotic diseases§New, re-emerging or drug-resistant infections whose incidence in humans has increased within the past two decades or whose incidence threatens to increase in the near future

Indonesia – A rapidly developing country with serious problems in Infectious diseaseMalaria:

15 million cases and 42,000 deaths/year (2005) - highest case number and fatality rate in the world; increasing drug resistant parasites

Tuberculosis:ranked third in TB burden following India and China - TB is third major causes of mortalityEstimation: 269 TB cases/100,000

Dengue: Most important viralborne disease2004: 78,690 cases (CFR- 1.2%)2007: 123,174 cases,1,251 deaths

Hepatitis B:10% of population are carriersModerate-to-high endemic (WHO)

Serotype Legend

4 13 2

42%

30%

8%

20%41%

32%

19%

8%

Makassar 2007-2008N = 111

Merauke 2001

Palembang 1998

Bandung 2002

Yogyakarta 1996

Jayapura 1994Jakarta 2004

Corwin 2001; Suwandono 2006; Porter 2005; Graham 1999; Richards 1997; Sukri 2003

DENV SEROTYPE DISTRIBUTION – SHOWED DIVERSITY

Four antigenically distinct serotypes: infection with one serotype does not provide protection to the other three

Management of Disease is Not Simple, Need a Strong Disease Surveillance

0

50000

100000

150000

200000

250000

1985 1986 1987 1988 1989 1990 1991 1992 1993 1994 1995 1996 1997 1998 1999 2000 2001 2002 2003 2004 2005 2006

C A

S E

Indonesia South-east Asia Region

SCIENCE & TECHNOLOGY IN HEALTH SECURITY –PREPAREDNESS FOR PANDEMIC

RISK ASSESSMENTS§ Molecular Epidemiology:

• Cluster of viral isolates will indicate the presence of new strain

• Surveillance - tracing sources of infection

§ Characteristics of Virus• Alteration of interaction with host

receptors - pandemic need changing in specificity of viral type receptor into human-type

• Change of virulence • Drug resistance

ALL ABOUT COLLABORATION

• Interdisciplinary collaboration between genetics, language, culture and medicine

Genetics

Linguistic

Anthropology Archeology

Medicine

Mathematic

THE AUSTRONESIAN DIASPORA

WMP

WMPCMP

PAPUA

OCEANIC

OCEANIC

SHWNG

§ One of the world largest language group. § Covers a very wide area, from Madagascar Island as the

western most to the Easter Island of Polynesia

EIJKMAN INSTITUTE and THE POPULATION STRUCTURE of the INDONESIAN ARCHIPELAGO

Eijk

man

Inst

itute

• More than 6,000 samples collected• More than 3745 individuals from 35 ethnic

populations examined for mtDNA• 2000 genetic samples from 25 isolated populations of

13 islands for Y STR • Most extensive database on mtDNA sequence

polymorphisms in Indonesian archipelago• Pan Asia SNP initiative – mapping 50K Asian

population using DNA microarray – 234 Indonesian samples

• 200 male samples of Javanese and Batak origin for Rapidly Mutating YSTR

• Forensic population database – STR marker 21 loci, Y filer

§ MtDNA and Y chr diversity of 2740 individuals, 70 communities, 12 islands§ Indonesian genetic diversity

is a direct outcome of complex history of immigration, transitory migrants and populationsTumonggor et al. J Hum Genet. 2013: 58

SETTLEMENT of ISLAND SOUTHEAST ASIATHE FIRST WAVE: FROM AFRICA TO THE EAST

~50,000 ya

Mellars P (2006). Science

The first stage of Indonesian prehistory represent the archipelago’s initial settlement as part of Africa dispersal ~ 50kya

SETTLEMENT OF ISLAND SOUTHEAST ASIATHE SECOND WAVE: THE AUSTRONESIAN EXPANSION

~6,000 ya

Austronesians

~50,000 ya

Taiwan

Madagascar

Polynesia

~8,000 ya

Neolithic movements into and around island SEA – may involve population dispersals from (into) Taiwan, and between Indonesian island groups

§ The largest survey of Indonesian Y chromosome showed the presence of multiple genetic strata that likely arose through a series of distinct migratory process § Paternal gene pool subdivided eastern and western part of

Indonesia

• 1928 individuals - 73 populations, 11 language families • Autosomal marker 50K - HUGO Pan-Asian SNP Consortium -.

Science 326, 1541 (2009)

• Autosomal data strongly support large demographic movements of Asian populations into eastern Indonesia from around 4 kya (Xu et al & HUGO Pan-Asian SNP Consortium, PNAS 109, 4574 (2012)

East Papuan and Alorese

Sumba, Flores, Lembata

Malay,Sumatra, Borneo, Java (Mix with Austroasiatic)

VARIATION OF GENOTYPIC PATTERN OF DISEASES

HEPATITIS AND OVALOCYTOSIS AS A MODEL

• Variation of ethnics in Indonesia: 1. The peopling of the Indonesian archipelago

using mitochondrial-DNA, Y and autosomal chromosome

2. Anthropological and linguistic study -Consistent with cultural and linguistic characteristics

• Variation of susceptibility and resistance to disease (malaria, TB and others)

• Variation of genotypic and phenotypic pattern of diseases (infectious and non-infectious, including hepatitis B/C, dengue,red blood cell disorders )

HEPATITIS B VIRUS GENETIC HETEROGENEITY

§ Ten HBV genotypes - A to J have been identified worldwide

§ Shows different geographical distribution, viral characteristics and possibly clinical outcomes and response to treatment, , provide historical information pattern of the local population

Miyakawa & Mizokami, Intervirology 2003;46:329-338

A: North-west Europe, North America, Central Africa; B: Southeast Asia; C: Far east; D: Mediteranean, Middle East, India; E: Sub-Saharan Africa; F: American natives, Polynesian; G.USA samples, H: Mexico

• Indonesian Archipelago:Genotypes B and C are predominant, with sub-genotype B3/adw unique to Indonesia

• Latin America:Genotypes F and H are indigenous in this continent, genotypes A and D might be a mere reflection of a past European migration, and genotypes B and C could represent a consequence of a recent Asian migration.

JapanKorea

Kajang, Makasar

Java

Padang

Nias & Mentawai

Dayak Benuaq

Sumba

Alor

East Flores

Chinese IndonesianBangkok

Mandar, Toraja

West Flores

Kotamobagu

TaiwanHongkong

Nanning Shanghai

JayapuraSentani

Merauke

Mataram

Philippines

Medan

Ternate, Ambon

East Asia

China

Japan

West IndonesiaPhilippines

Nusa Tenggara islands of Indonesia

C1

C2

B2B3B5B7

B9B8

D1

B1

C5

A1

Philippines

D3

HEPATITIS B VIRUS GENOTYPE DISTRIBUTION IN INDONESIAN ARCHIPELAGO

AY226578M57663A1AB246335A1AB241115A1AF297621A1

X70185A2AB222707A2AB126580A2

AB246337A2 DQ463787B6DQ463790B6

DQ463791B6DQ463792B6DQ463788B6

DQ463793B6DQ463794B6

DQ463789B6D23678B1AB073850B1AB073848B1D00329B1

DQ993700B2EU139543B2

DQ993708B2EIH21AB22061B2DQ993711B2EIH45AB2

AP011084B2 AP011089B7AB493833

AB493835AB493836

AB493832EIHB134B3AB033555B3

AP011085B3AB033554B3

D00331B32059B3

1839B3M54923B3

LBY01B7LBY041B7Alr049B7

AP011090B7LBY009B7

AP011091B7FLT32B7

AP011092B7KD048B7

CBL027B7LAR070B7

AP011088B7FLT020B7

AB493830AB493828

AB493829AB493827

AB493831.KDI35B8KDI43B8

KDI04B8KD35B8

AB219426B5AB219429B5

AB219427B5AB219428B5

AP011087B5AP011086B5

PAN09B9PAN011B9

Alo36B9PAN01B9PAN037B9

AP011096B8AB493834

AP011093B8AP011095B8

AP011094B8AY033073B4

AY033072B4AB100695B4

AB073835B4AB112066C1AB112348C1

AB112471C1AB074756C1

EIHB006C1AP011097C1SLK-126

AB033553C2AB113879C2

AB202071C2AP011098C2

X01587C2PhLC14C5

PhLC03C5AB241112C5

AP011101C5AP011099C5AP011100C5

PhCH24C5M154AP011105C7AP011104C7

AP011107C7AP011106C7

AP011108CJ022

STN013AB493837AB493847AB493838AB493840

AB493839AP011102C6AB105172

AB493844AB493843

AB493842AB493841

AP011103C6M007X75656C3

X75665C3AB048705C4

AB048704C4AF280817D1

AY161157D1AY161150D1

M73D1AF151735D1

AB078032D2AB078033D2

AB090268D2AB090269D2AY090452D3

AB493848.AB493845

AB493846DQ315776D3

DQ315777D3AB048702D4

AB048703D4AB048701D4AB033559D4

AB032431EAB205129E

AB106564EX75657E

AB064311GAB064312GEF634480G

AB056513GAB086397F1

AF223963F1AF223962F2AF223965F2

AB166850F2AB214516F2

AY090455F1AB179747H

AB205010HAB266536HAB059660H

100

100

100

100

100100

100

100

100

99

100

100100

100

100

100

100

100

100

100

100

100

100

100

100

100

92

100

92

100

92

100

100

68

100

100

92

92

96

92

100

100100

92

100

92

100

100

97

100

100

100

100

100

100

0.02

100

HF

HF2

F1GG

ED

D1

D2

D3D4

E

C3C4

C6

C7

C5

C2

C1

B4

B8

B5

B9

B7

B3

B2

B1

B6

A1A2

C

B

AInuit populations in Arctic

Japanese, east Asia

Chinese, Asia mainland

Indonesian populations, Southeast Asia

Indonesian populations, Southeast Asia

Indonesian populations, Southeast AsiaPhilippino and Indonesian populations, Southeast Asia

Indonesian populations, Southeast Asia

Vietnamese, Southeast Asia East Asia, China mainland and Southeast Asia

East Asia, China mainland and Southeast Asia

Philippino, East Asia

Indonesian populations, Southeast Asia

Indonesian populations, Southeast Asia

Polynesia, New Caledonia, Pacific region Aborigine populations, Southern Australia

Figure 2

Balinese and LombokSumbanese

Flores

Alorese

Minahasa and Talaud Ternate and Ambonese

C1

C5

BatakB2

B3B7B9

C2

Malay C2

B3B9

C1B2

B3

B8B9

C1C2

B3

B7B5

B8B9

B3B5B7B9

C2C1 Papuan

C1C2C6

B3B7

Indonesian of Chinese ethnic origin

B2B3B5

C1C2

B3B5

B8B9

Nias and Mentawai

Minang

B3

C5

B7

bMakasarese

C1C2

B2B3B5B8B9

C1

C2

Javanese

B5B3

B7

C1

* Indonesian of Chinese ethnic origin

*

a

B7B5

B8

B9

C1C2

Torajan and Mandar B3

B7

C1C2

B5B3B5B7B9

Javanese(Suriname)

Javanese(Holland)

Javanese (Java)

HBV/B3 (adw)

HBV genotype is maintained in Javanese ethnic population separated for centuries

100

AB540583

AF223957C1

AP011097

AF223960AB112471

AB074756

AF068756

AB112348AB112066

AY247031

AF533983

X01587

D23681

AB033553

AB113879

AP011098

AB202071

PhCH24AB241112

PhLC03

PhLC14 AP011100

AP011101

AP011099AP011106

AP011107

AP011104

AP011105

EU670263AP011108

GQ358157

AB493837

GQ358155

GQ358156

AB493840 AB493838AB493842

AB493844

AP011103

AB493841

AB493839

AP011102

X75656

X75665

AB048705

AB048704

AB266536

6

3

4

97

8

5

21

10

H

EAST ASIA AND SEA

Papua-Pacific

North Australia

TWO TYPES HBV/C SUBGENOTYPES: THE ASIAN AND PAPUA-PACIFIC(study on HBV isolates from the Asia-Pacific Region)

• Subgenotypes of HBV/B in Japan (Sugauchi et al):– Bj

– B2 (Ba: prevalent in China)

Maintained Viral Characteristics

In Chinese ethnic population*

Clinical Implications?

DNA Sequence:

*Have been living in Indonesia for > 3

generations

HBV genotype is maintained in Javaneseethnic population separated for centuries

• Subgenotypes of HBV/B in Indonesia:– B3 (adw)– B5 (adw)– B7 (ayw)– B2 (Chinese

Indonesian)

‘a’ determinant (124-147)

309 regular blood donors (2005) from Medan (North Sumatra) and Solo (Central Java) (HBsAg, anti-HCV and anti-HIV neg)• anti-HBc (+) : 134 (43.4%)• HBV DNA (+): 25 (8.09%)• Variants were detected in 7 samples: T123A (1), M133L (1),

T143M (7)

Thedja MD et al. 2010. Occult hepatitis B in blood donors in Indonesia: alter antigenicity of the hepatitis B virus surface protein. Hepatology Int.4, 608

CLINICAL AND PUBLIC HEALTH SIGNIFICANCE:Detection failure of HepB virus in blood donors

• Malaria - caused by Plasmodium parasites- is responsible for high mortality mortality. An estimated 300-500 million cases each year result in more than 1 million deaths

• Available preventive methods are not sufficient• Humans have a number of genetic adaptations

that act to combat Plasmodium

• Understanding the mechanism of malaria interactions with the erythrocytes membrane will provide opportunities for new methods of disease prevention and treatment

Rank Country Mortality Rate

49 INDONESIA 3.1

MALARIA AS A SELECTIVE AGENT IN HUMANS

WHO 2005

SAO, HbE,Gerbich negativity,

α-thalassemia (many), G6PD deficiency (many)

Duffy negativity,G6PD deficiency (A-),

HbS, HbC

COMMON THEMES AMONG MALARIA-RESISTANCE ALLELES EXAMINED to DATE

(G6PD A-, HBC, HBS, DUFFY NEGATIVITY, HBE)

1) Recent origins (<<30,000 years)2) Strong selection coefficients3) Compatible with “Malarial Eve” Hypothesis

• An uncommon variant of hereditary elliptocytosis• Caused by heterozygosity for a 27 bp del in the gene

encoding the erythrocyte membrane protein band 3 – the solute carrier family 4 (SLC4A1) protein on chromosome 17

• SAO erythrocytes are rigid because mutation enhances the tightness of association between band 3, ankyrin and the spectrin lattice

• The condition confers highly specific protection against cerebral malaria

SOUTHEAST ASIA OVALOCYTOSIS

§ Erythrocyte membrane contain variety of proteins –(i) peripheral proteins of bands 1, 2, 4.1, 4.2, 5 and 6, and (ii) integral proteins of bands 3, 7 and PAS 1 - 4.

§ Mutation in SLC4A1 gene enhances the tightness of association between band 3, ankyrin and spectrin lattice

SAO PROTECTS FROM MALARIA-RELATED MORTALITY.

• Strong correlation between SAO and malaria prevalence (Mgone et al. 1996).

• Case-control studies suggest strong (complete?) protection from cerebral malaria (Genton et al. 1995, Allen et al. 1999).

SAO is A BALANCED POLYMORPHISM§ Heterozygote has greatly enhanced fitness

in malarial environments.Heterozygotes have no negative clinical manifestations.

§ SAO homozygote is inviableWhen SAO is common, it can demonstrably increase the miscarriage rate (Liu et al. 1994)

§ SAO as a health burdenOne in for of the pregnancies in these couples will be lost due to SAO inviability.With random mating 12.25% (=0.35x0.35) of couples will both be heterozygous carriers of SAO.

SAO HAS A RESTRICTED GEOGRAPHIC DISTRIBUTION.

GEOGRAPHICAL DISTRIBUTION of SAO in INDONESIAN ARCHIPELAGO (as 2014)

0

8.6

0

11 01.4

2.9 010.5 13.4

27.20

2.6

0

06.6

1.8

4.6

4 3.3

0.9

20

30

4.6

Eijkman Institute

1.4

8.6

1.8

3.3

4.6

0.9

6.6

10.5

2.9

27

13.4

0

0

0

0

0

0

0

0

0 10 20 30

0 . 1

A f r i c a

A l o r

I r i a n

B a t a k

J a v a

M a k a s s a r

S a s a k

B u g i s

M i n a h a s a

S u m b a

S u m b a w a

T o r a j a

K a i l i

B a n j a r

P a l e m b a n g

T e n g g e r

D a y a k

M i n a n g

M a l a y ( P ’ b a r u )

B a l i

FREQUENCY OF SAO (%)

FREQUENCY OF SAO VARIES SIGNIFICANTLY IN DIFFERENT POPULATIONS (not revised)

DISTRIBUTION OF PAPUAN GENETIC CONTRIBUTION AND SAO PREVALENCE

Papuan Genetic Contribution (%)0 5 10 15 20 25 30

SAO

Pre

vala

nce

(%)

0

10

20

30

40

50

§ SAO is not found in non-endemic area for malaria; but not all endemic area shown SAO

•SAO is found mainly in two clades: a. Malay-related (lowprevalence) and b. Papuan-related (high prevalence)

SAO likely evolved after expansion of P. falciparum into Southeast Asia.

P. falciparum emerges as health threat ~10,000 years before present.

Models of Coevolution between Plasmodium and Humans May Need to Accommodate the Existence of Ancient Malaria-resistance Alleles

FUTURE STUDY§ Bringing human and medical genetics and

computational biology together – to answer the big question on how culture shaped the human genome§ The answer is in your hands

Genetics

Linguistic

Anthropology Archeology

Medicine

Mathematic

Acknowledgment

Steve Lansing, Complex Institute, NTUMurray Cox and Elsa Guillot, Massey University, New ZealandTatyana Karafet, University of ArizonaThe HUGO Pan-Asian SNP Consortium

EIJKMAN INSTITUTE

David Mulyono,Meta Dewi TedjaDin Syafruddin

Helen SuryadiSafarina MalikGludhug PurnomoWindy JoanmawatiAlida HarahapDewi Megawati

Thank you and greetings from Indonesia