Gastroenteritis Viruses

(VIRUS-VIRUS
PENYEBAB DIARE)
A. T. AMAN, MD., PhD.
Clinical Microbiologist

DEPT. OF MICROBIOLOGY
FAC. OF MEDICINE
UGM, YOGYAKARTA
Blok Nutrisi dan Sistem Digesti, 28 April 2008
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References
1.

2.
3.

Richman D. D. et al (ed). Clinical

Virology. 2nd ed. ASM press,
Herndon, VA, USA
Murray et al., Manual of Clinical
Mycrobiology, 8th ed. (2003).
Brooks et al. Jawetz, Melnick, and
Adelbergs Medical Microbiology. 23rd
ed. 2004.
2

Termasuk disini:
1. Rotavirus.
2. Norwalk-Like virus
3. Sapporo viruses
4. Astroviruses.
5. Enteric Adenovirus

Introduction
Gastroenteritis Viruses:
Kuman pathogen penting, mrpk group luas,
meliputi Family:
-. Reoviridae: rotavirus
-. Adenoviridae: Enteric adenoviruses
-. Caliciviridae: Norwalk-Like viruses,
Caliciviruses.
-. Astroviridae: human astroviruses.
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Introduction (cont,d)
Mengapa Penting:
1. Penyakit Diare di neg. sdg berkembang diperkirakan:

-. Pd anak < 5 th: 1 milyard/th.

-. 3.3 million death.
2. Sebagian besar tdk diketahui penyebabnya.
3. Th 1973: Rotaviruses was discovered (by EM).
4. Rotaviruses: The most important cause of severe
diarrhea in infants and children <5 yrs, in
developed & developing countries worldwide.
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I. ROTAVIRUS

Epidemiology of Rotavirus (Grup A)

1. RV adalah penyebab utama:
Diare berat.
Terutama pd anak <2 th
Baik di neg. industri maupun neg. sdg berkembang.

2. Di USA:

RV penyebab: 5-10% diare pd anak balita.

Penyebab 30-50% episode diarrhea berat.
Estimated: 2.7 million episodes per tahun.
50, 000 hospitalizations pertahun.

3. Perkiraan di neg. sdg berkembang (1997):

- 130 million, dg 18 juta, diare moderate-berat.
- Kematian 873, 000 infants dan anak balita.

An Estimate of the role Rotavirus in Causing

Severe Diarrhea (in children <2yrs) in Developed &
Developing Countries (Kapikian, JAMA, 1993)

Estimated global distribution of the 800,000 annual

deaths caused by rotavirus diarrhea

Epidemiology of Rotavirus
Indonesia
Rotavirus di Ind. (< 5 th):
Di Yogyakarta: June 78- June 79), dg ME:
38% diare akut (Soenarto et al., 1981).
Studi di berbagai RS di Jakarta, dg EIA: 42%
diare akut (Subekti et al., 2002).

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Seasonal Pattern
Infeksi Rotavirus di pengaruhi musim.
*. Di negara dg iklim dingin: puncak
pd musim dingin.
*. Penyebab: diduga faktor
kelembaban yg rendah menaikan
survival virus.
*. Di neg. sdg berkembang/iklim
tropik: sebag. Data menunjukkan tdk
ada pola musiman
Tetapi data terakhir di Indonesia: ?.
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Rotavirus: General Features

1. Ditemukan oleh Bishop, 1973 (dg ME).
2. Virion: like a wheel dg duri pendek &
outer rim halus (rota =wheel).
3. Diameter: 70 nm.
4. Dua lapisan protein capsid: lapisan luar
dan dalam.
5. Lapisan berikutnya: protein core.
6. Di dlm core: genom, berupa 11 dsRNA.
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Rotavirus (by EM) in stool filtrate

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Genom Rotavirus
1. 11 segment of dsRNA.
2. Ukuran: 0.6 to 3.3 Kb.
3. Berdasar ukuran genom, dibagi mjd: 4
klas.
4. Ke 11 segmen mengkode:
6 protein structural, Viral proteins (VP).
Segment 1-3 mengkode core & subcore: VP1,
VP2 and VP3.
6 protein non-structural : NSP1 to NSP6.

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Antigenic Composition
RV memp. 3 antigenic specificities: group,
subgroup and serotypes.
Yg tlh temukan: Group A, B, C, D, E, F & G.
Group A: pemyebab utama diare pd <5 th.
- Group: B outbreak di Cina.
- Group C: diarrhea in India.

* Subgroup: memakai VP6, digunakan sbg

epidemiological marker
*. Serotype pd group A, diidentifikasi dg:
Neutralization assay
RT-PCR.

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Rotavirus: virion structure

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Serotyping
Berdasar VP7: serotype G (glycoprotein).
Berdasar VP4: serotipe P (protease
sensitive).
Dominant G serotype di dunia: G1-4 &
G9.
Antibody thd G1-4: ada reaksi silang.
Serotipe tertentu dominan di neg./daerah
tertentu.
Serotipe P yg dominan: P2, P4, P8.
Yg dominan di Indonesia?.
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Serotipe Rotavirus di Indonesia

Dg antibodi monoclonal (Bishop et al, 89):

59 of 111 samples (53%) can be type.
Serotype 1: 1.6 (.%) ; Type 2: 5 (8.5%);
Type 3: 31 (52.5%) and Type 4: 21 (35.6 %).

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G-type Comparison: data 78-79 and 01-03

Year
data

No.
test.

Type (%)
G1

G2

78-79 111

1
(1.6)

01-03 104

61 11
(59) (10)

G3

5
31
(8.5) (52.5)

Method

G4

Un-typ.

21
(35.6).

52 (47)

Mon.
Ig.

16
(15)

7 (7)

RTPCR
+ Hyb

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Gambaran Klinis
1. Inkubasi: 1-4 hari.
2. respon thd Infeksi RV infection
bervariasi: mulai dari subclinicas, diare
ringan s/d berat bahkan dpt mengakibatkan
kematian.
3. Gambaran utama:

Demam (380C or above).

Diarrhea cair.
Dehidrasi.
Muntah.
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Clinical Features (cont,d)

4. Biasanya: berat pd infant & anak
balita, tetapi kurang berat pd
neonatus dan dewasa> Why ?.
5. Lama gejala: 4-5 hari.
6. Virus shedding: 6-10 hari.

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Pathogenesis and Pathology

1. Lesi akibat infeksi RV:
Pd mukosa jejunum mucosa: pemendekan
dan atrophy villi.
Irregular microvilli.
Infiltrasi Mononuclear pd lamina propria.

2. Mekanisa tjd diare: belum banyak

diketahui, diduga berkaitan dg malabsorbtion.
3. Kemungkinan mulai dengan:
perubahan morphologi (e.g.: flattening
of epithelial cells) pd bag. Atas usus
halus.
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Pathogenesis & Pathology

Percobaan bnt (calves): diberi human
rotavirus, berakibat:
-. Dlm 30 mnts: pemendekan villi &
pendataran sel epithel pd bag. atas
usus halus tetapi antigen RV tdk
ditemukan.
- Dlm 30 mnts; tdk tjd perubahan bag.
Bawah usus halus, tetapi antigen RV
sanghat banyak.
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Pathogenesis and Pathology

-. Dlm 7 jam: tjd perubahan morphologi
pd bag. Bawah usus halus.
40 jam sth ,ulai diare, usu kembali
normal.
Terjadi ganguan transport glucose
danSodium.
-. Tdk ada bukti keterlibatan
adenylate cyclase m/p cAMP
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IMMUNITAS
Mechanisme immunitas terhadap RV masih
sedikit yg diketahui
Newborn calves akan menderita diare, meski
memp. circulating antibodies sedang-tinggi dari
colostrum. > ..>circulating antibodies tidak
mampu memproteksi.
Data lain: Ig pd lumen usus merupakan
protektor utama.
Jika circulating antibodies cukup, mk supply
thd antibodi lokal akan memberikan proteksi.
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IMMUNITY (contd)
Which Antibodies involve?
Adaptive: cellular ?,
hummoral ?, Ig ..?.

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IMMUNITY (contd)
Data penelitian:.
RV specific fecal IgA dg konsentrasi
1/80 /min. 1/20 terkait dg protecsi thd
infeksi atau manifestasinya/sakit.
Titer anti-RV serum:
IgA > 1/200 atau IgG > 1/800 memberi
perlindungan thd infeksi.

Jika IgA tdk ada, maka antibodi lain,

spt: IgG akan memberi proteksi thd
infeksi yg homolog.
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IMMUNITY (contd)
Peran cell-mediated immunity:
RV specific antibodi penting utk protekasi
thd reinfection.
Homologous RV strains jauh lebih kuat
drpd heterologous RV strains dlm
menginduksi humoral immune response
lokal.
MHC class I-restricted CD8+ T cells:
berpernan palingh besar dlm
menyembuhkan infection primer, tetapi
CD4+T cells juga terlibat.
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IMMUNITY (lanjutand)
Peranan cell-mediated immunity:
-. CD8+T cells pd proteksi thd reinfection:
Lengkap s/d: 2 mg.
Partial: stlh 3 bulan.
Hampir hilang stlh 8 bl.

.> Why the incident is very low in > 5

years of age ? (explain!).
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DIAGNOSIS
Melihat manifestasi Klinis akibat infeksi RV.
Deteksi virus atao antigennya atau respon
antibodi.
Deteksi Virus:
EM, highly specific (relatively less
sensitive)
Dg RT-PCR.
Deteksi antigen pd feses: ELISA, highly
specific.
Deteksi antibody (IgA and IgG): ELISA.

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TREATMENT
Primary treatment: Rehidrasi, dg
terapi standard WHO/Indonesia.

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Pencegahan
Tujuan: mencegah diare berat pada 2
th pertama.
Infection alami tdk memberikan
perlindungan yg lengkap.
Local intestinal immunity perperanan
penting.
Pencegahan: vaksinasi utk
merangsang IgA dan antibodi lokal
lainnya.
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II. Norwalk-Like virus

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Introduction
Human Caliciviruses dibagi
mjd (berdasarkan sequence &
gambaran morphologic):
a. Norwalk-Like virus
b. Sapporo viruses

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Introduction (lanjutan)
Norwalk-Like Viruses:
1.
2.
3.
4.
5.

The first viral agents to be associated

with Gastroenteritis.
Discovered in outbreak in Norwalk,
Ohio (1972) by immune EM.
Have cause outbreak: Cruise ship,
restaurants and in family setting.
Includes: Hawaii, Snow Mountain &
Taunton viruses.
The major cause of non bacterial
epidemic gastroenteritis.
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Structure and genome

Norwalk-Like Viruses:
-

Round, diameter 27 nm, permukaan

luar kasar.
Non-envelop viruses.
Genome: ssRNA, ukuran 7,7 kb, post. Sense,
mgd 3 major ORFs utama,
ORFs: mengkode: capsid, helicase, RNA
dependent RNA pol.

Ditemukan pd feces patients

gastroenteritis.
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Other characteristic
Norwalk-Like Viruses:
-

Tetap infective dlm keadaan:

- pemaparan stool filtrate thd pH 2.7, 3 jam, RT.
- treatment dg 20% ether, pd 40C, 18 jam.
- treatment dg 3.75 -6.26 mg/L chlorine (kadar
yg ditemukan pd sistem distribusi air minum).
..> Inactiv dg: 10mg/L chlorine (konsent. Utk
tretment air stl kontaminasi.

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Clinical Features
1. Incubation period: 12-48 hrs
2. Infeksi dpt berakibat: asymptomatic-diarrhea
diare berat.
3. Gambaran utama: muntah dan atau diare,
mendadak, tanpa tanda prodromal.
3. Gambaran klinis utama meliputi:

Nausea and vomiting.

Watery diarrhea.
Dehydration.
Abdominal cramp.
Headache and fever.
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Diagnosis lab.
1. Gambaran Klini + tdk ditemukan
penyebab parasit, bakteri.
2. DG EM (tdk begitu sensitif). Yg
terbanyak dg IEM, dan masih terbatas utk
riset.
IEM: dpt deteksi antibody dan antigen.

3. Kultur: belum dpt dikembangkan.

4. Teknik molekular: RT-PCR, paling
sensitif.
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III. Sapporo viruses

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Introduction
1.

The first discovered from stool of

an asymptomatic patient.

2.

Pathogen ?. In an outbreak of
gastroenteritis, the virus was found
in the stool followed by
seroconversion.

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Virus characteristic
Sapporo Virus:
-

Retain infectivity after:

- exposure of stool filtrate to pH 2.7 for 3 hrs at
RT.
- treatment with 20% ether, at 40C, 18 hrs
- treatment with 3.75 -6.26 mg/L of chlorine
(cost. Found in drinking water distribution
system). Inactivated with 10mg/L of chlorine
(conct. To treat water after contamination).
-

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Clinical Features
1. Incubation period: 48-72 hrs
2. Infection can produce: asymptomaticsevere diarrhea, 1-11 days.
3. Mainly affected infant young children.
4. Occurs: throughout the year, peak in
winter.
5. Transmission: food and water.
6. Distribution: world-wide, all pop. is
Seropositive by the age of 4 year.
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IV. Astroviruses.

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Introduction
1.

The first discovered by EM, from

stool of patient with gastroenteritis
(1975).

2.

Human astroviruses: 8 serotypes,

with serotype 1 is the most
prevalent.

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Clinical Features
1. Incubation period: about 3-4 days
2. Infection can produce:
asymptomatic-diarrhea, 3-14 days.
3. Mainly affected infant young
children.
4. Neutralizing antibody: by 1 year.
5. Antibody to serotype 1 is the most
prevalent.
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V. Enteric Adenovirus
1. Infection can cause prolong diarrhea,
with low grade fever, with or without
vomiting.
2. Lasted up to 14 days.
3. Considered to the etiologic agent if
found abundant in patient with GE.
4. Common cause of pediatric GE, with
peak under 2 yrs.
5. Infection occurs all year around.

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Diagnosis
1. Specimen collection and Storage:
- stool should be collected as soon as
possible, ideally within 48 hrs.
- Rectal swab: not recommended, not
enough.
- Sent without transport medium
- Storage at 40C (up to a week)
- Storage for prolong: -700C.

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Diagnosis
2. Virus Detection:
-. Direct examination by EM
-. IEM.
-. Antigen detection assay: ELISA, Latex
Aglutination
-. Assay based on the viral genome:
-. RT-PCR.
-. Identification of dsRNA by RNA PAGE.
-. Dot Blot Assay.

3. Interpretation: ?.
- What factors should be considered ?.
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THANK YOU
VERY MUCH

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Structure and genome

Sapporo Virus:
-

Round, with 35 nm in diameter,

with well define outer edge.

Genome: ssRNA, post. Sense, 8 kb

containing 2 ORFs.
- Main protein being codes: capsid.

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