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UNIVERSITI PUTRA MALAYSIA

PREVALENCE OF AND FACTORS AFFECTING ACQUISITION OF INTESTINAL CRYPTOSPORIDIOSIS IN MALAYSIAN CHILDREN WITH MALIGNANCIES AT ONCOLOGY WARD, INSTITUTE OF PEDIATRICS,

HOSPITAL KUALA LUMPUR, MALAYSIA

LUBNA MOHAMED ELBESHTI

FPSK(m) 2011 42

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PREVALENCE OF AND FACTORS AFFECTING ACQUISITION OF INTESTINAL

CRYPTOSPORIDIOSIS IN MALAYSIAN CHILDREN WITH MALIGNANCIES AT

ONCOLOGY WARD, INSTITUTE OF PEDIATRICS, HOSPITAL KUALA LUMPUR,

MALAYSIA

By

LUBNA MOHAMED ELBESHTI

Thesis Submitted to the School of Graduate Students, University Putra Malaysia, in Fulfillment

of the Requirements for the Degree of Master of Science

June 2011

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Abstract of thesis presented to the Senate of Universiti Putra Malaysia in fulfillment of the

requirement for the degree of Master of Science

PREVALENCE OF AND FACTORS AFFECTING ACQUISITION OF INTESTINAL

CRYPTOSPORIDIOSIS IN MALAYSIAN CHILDREN WITH MALIGNANCIES AT

ONCOLOGY WARD, INSTITUTE OF PAEDIATRICS, HOSPITAL KUALA LUMPUR,

MALAYSIA

By

LUBNA MOHAMED ELBESHTI

June 2011

Supervisor: Rukman Awang Hamat, PhD

Faculty Medicine and Health Sciences

Cryptosporidium parvum is an opportunistic parasitic agent that has a world-wide distribution.

The disease caused by this parasite can be severe and very difficult to manage in

immunocompromised patients especially in children with malignancies. However, data on

immunocompromised children in Malaysia is very much lacking. The aim of this study was to

estimate the prevalence of cryptosporidiosis in children with different types of malignancies and

to study factors that might contribute to acquisition of cryptosporidiosis.

A cross-sectional study was conducted over 10 months from November 2009 until August 2010

in Institute of Pediatrics, Hospital Kuala Lumpur. A self-administered questionnaire was used

and medical records were obtained. Stool samples were examined for the Cryptosporidium

oocyst by using two different techniques i.e. modified Ziehl-Neelsen stain and

Immunochromatographic (ICT) assays (RIDA-Quick Cryptosporidium R-Biopharm, Germany).

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One hundred and ten stool samples were collected from children (56 boys, 49 girls) with

different types of malignancies between the ages of 3 months and 17 years (mean age: 2 years).

The majority of those children were the Malays (75.2%), followed by the Chinese (11.4%),

Indians (8.6%) and others (4.8%) The most common type of lympho-hematopoietic malignancies

was acute lymphoblastic leukemia (38.1%), followed by acute myeloid leukemia (8.6%),

suspected leukemia (8.6%), lymphoma (7.6%), and chronic myeloid leukemia (1.9%). Whereas,

among non-lympho-hematopoietic malignancies, brain tumor represented 11.4% of cases,

followed by retinoblastoma (5.7%), hepatoblastoma (3.8%), Wilm‟s tumor (2.9%),

pleuropulmonary blastoma (1.9%) and right adrenal cortical tumor (0.9%). Fever was presented

in (83.8%) of those patients, followed by diarrhea (54.3%). All stool samples were negative for

Cryptosporidium oocysts by two different techniques. (33.3%) from those patients had history of

admission to other wards, (29.5%) had history of animal contact, (24.8%) had history of

swimming in the swimming pool. In terms of precautionary measures practiced, (80.9%) and

(75.2%) washed their hands before and after eating, or after going to the toilet respectively. In

addition, preventive measures that were also observed: (16.2%) had history of admission to day-

care center, (2.9%) had history of drinking tap water, and (0.9%) had history of travel.

In this present study, we documented a zero prevalence rate of cryptosporidiosis amongst

children with malignancies despite higher prevalence rates being reported in other developing

countries. Our results may suggest that the children with malignancies are at low risk of

acquiring cryptosporidiosis because of good personal hygiene, good infection control and

practices in the hospital, and improve water supply system. Screening for Cryptosporidium

oocysts may not be necessary in those patients. Our findings suggest that routine screening of

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stools for Cryptosporidium might not be necessary for our patients. However, we believe large

multicenter studies throughout Malaysia are needed to establish the true prevalence and

characteristics of cryptosporidiosis among children with malignancies.

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Abstrak projek yang dikemukakan kepada Senat Universiti Putra Malaysia

Sebagai memenuhi keperluan untuk ljazah Master Sains

PREVALENS DAN FAKTOR YANG MEMPENGARUHI KEJADIAN

KRIPTOSPORIDIOSIS USU PADA KANAK-KANAK MALAYSIA YANG

MENGHIDAPI MALIGNANSI DI WAD ONKOLOGI , INSTITUT PEDIATRIK,

HOSPITAL KUALA LUMPUR , MALAYSIA

Oleh

LUBNA MOHAMED ELBESHTI

Jun 2011

Pengerusi: Rukman Awang Hamat, PhD

Fakulti Perubatan dan Sains Kesihatan

Cryptosporidium parvum adalah sejenis agen parasit oportunistik yang mempunyai taburan di

serata dunia. Penyakit yang di sebabkan oleh parasit ini boleh menjadi parah dan ia sangat sukar

untuk di kawal pada pesakit terimunokompromi terutamanya kanak-kanak malignansi. Namun

begitu, data pesakit terimunokompromi di Malaysia tidak mencukupi. Tujuan penyelidikan ini di

jalankan adalah untuk menganggarkan prevalen cryptosporidiosis pada kanak-kanak yang

mempunyai malignansi berlainan dan mengkaji factor-faktor jangkitan cryptosporidiosis.

Satu kajian keratan rentas telah di jalankan selama 10 bulan dari November 2009 sehingga Ogos

2010 di Institut Pediatrik, Hospital Kuala Lumpur. Soal selidik yang di jalankan sendiri di

gunakan dan rekod perubatan di perolehi. Sampel tinja telah di periksa untuk kehadiran oosista

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Cryptosporidium menggunakan 2 teknik berlainan iaitu pewarna Ziehl-Neelsen terubahsuai dan

asai imunokromatografi (ICT) (RIDA-Quick Cryptosporidium R-Biopharm, Germany).

Seratus dan sepuluh sampel tinja telah di kumpul dari kanak-kanak (56 lelaki, 49 perempuan)

dengan malignansi yang berlainan, yang berumur di antara 3 bulan dan 17 tahun (min umur: 2

tahun). Sebahagian besar daripada kanak-kanak tersebut adalah Melayu (75.2%), diikuti dengan

Cina (11.4%), India (8.6%) dan lain-lain (4.8%). Malignansi berjenis hematopoetik limfa yang

paling umum adalah leukemia limfoblastik akut (38.1%), di ikuti dengan leukemia mieloid akut

(8.6%), pesakit yang disyaki leukemia (8.6%), limfoma (7.6%), leukemia mieloid kronik (1.9%).

Manakala di kalangan malignansi yang bukan berjenis hematopoetik limfa, kes tumor otak

mewakili 11.4%, diikuti dengan retinoblastoma (5.7%), hepatoblastoma (3.8%), tumor Wilm

(2.9%), blastoma pleuropulmonari (1.9%) dan tumor korteks adrenal kanan (0.9%). Lapan puluh

tiga perpuluhan lapan peratus pesakit mengalami demam, diikuti dengan cirit-birit (54.3%).

Semua sampel tinja di dapati negatif untuk oosista Cyrptosporidium berdasarkan 2 teknik

berbeza. Tiga puluh tiga dan tiga peratus daripada pesakit tersebut mempunyai sejarah

kemasukan ke wad berlainan, (29.5%) mempunyai sejarah menyentuh binatang, (24.8%)

mempunyai sejarah berenang di dalam kolam air. Berdasarkan langkah berjaga-jaga yang di

amalkan (80.9%) dan (75.2%) mencuci tangan sebelum dan selepas makan, serta selepas

mengunakan tandas. Di samping itu, langkah berjaga-jaga tersebut telah di patuhi: (16.2%)

mempunyai sejarah menghadiri pusat asuhan kanak-kanak, (2.9%) mempunyai sejarah minum

air paip, dan (0.9%) mempunyai sejarah perjalanan.

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Berdasarkan kajian yang telah dijalankan, kami telah mendokumenkan kadar prevalens sifar

cryptosporidiosis di kalangan kanak-kanak malignansi walaupun negara membangun lain telah

melaporkan kadar prevalen yang tinggi. Keputusan kami mungkin menunjukkan bahawa kanak-

kanak malignansi mempunyai risiko yang rendah untuk di jangkiti cryptosporidiosis kerana

kebersihan diri yang terjaga, kawalan dan penjagaan infeksi yang baik di hospital serta sumber

air yang bertambah baik. Saringan terhadap oosista Cyptosporidium mungkin tidak diperlukan

bagi pesakit kita. Walau bagaimanapun, kami percaya yang keputusan kami mewajarkan kajian

lanjut seperti kajian prospektif longitudinal kawalan kes di masa hadapan. Kajian ini mungkin

dapat menolong kami menentukan status imunologi kanak-kanak yang menghidap cancer sama

ada mereka lebih cenderung mendapat cyrptosporidiosis.

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ACKNOWLEDGEMENTS

By the Name of Allah, Most Gracious, Most Merciful, all praise and thanks are due to Allah, and

peace and blessings be upon His Messenger. I would like to express the sincerest appreciation to

those who made this work possible: Advisory members, Friends and Family.

I would like to convey my deepest and grateful thanks to my supervisor, Dr. Rukman Awang

Hamat for providing me the opportunity to complete my study under his valuable guidance, for

the many useful advice and discussions, for his constant encouragement and guidance. I would

like to thank all the co-supervisors, Dr. Amal .R. Nimir, Dr. Ngah Zasmy Unyah. I am grateful

for their willingness to serve on my supervisory committee, constant encouragement, helpful

advice and many fruitful discussions. In addition, I would like to thank Dr. Malina Osman for

her help in the statistical analysis of the data.

Special thanks go to the staffs in Oncology Ward, Pediatric Institute, Hospital Kuala Lumpur,

who had given me a lot of helping hands during data collection. I would also thank Staffs at

Laboratory of Parasitology, UKM, especially Encik Ismail Hj Mohd Ghauth for his assistance

for doing the stain technique. Laboratory assistants of mainly Cik Nurhanim Kamaruddin and

Encik Mohd Nawawi Daud from Parasitology Laboratory, Faculty of Medicine and Health

Sciences, UPM.

I would also like to give all my respect and appreciation for all respondents of this study for

their cooperation all along the data collection.

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The People‟s Bureau of the Great Socialist People Libyan Arab Jamahiriya is gratefully

acknowledged for providing the financial support. Trust Account no: 6381600. .

Last but not least, very special thanks to my husband, his love, support and encouragements are

behind my success.

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APPROVAL SHEET

I certify that an Examination Committee has met on (14 June) to conduct the final examination

of Lubna Mohamed Elbeshti in her thesis entitled " Prevalence of and Factors Affecting

Acquisition of Intestinal Cryptosporidiosis in Malaysian Children with Malignancies at

Oncology Ward, Institute of Pediatrics, Hospital Kuala Lumpur, Malaysia " in accordance with

Universities and University Colleges Act 1971 and the Constitution of the Universiti Putra

Malaysia [P.U.(A) 106] 15 March 1998. The committee recommends that the student be awarded

the degree of Master of Science.

Members of the Thesis Examination Committee were as follows:

Zamberi Sekawi, PhD

Associate Professor

Faculty of Medicine and Health Sciences

Universiti Putra Malaysia

(Chairman)

Wan Omar bin Abdullah, PhD

Professor

Faculty of Medicine and Health Sciences

Universiti Putra Malaysia

(Internal Examiner)

Malina binti Osman, PhD

Lecturer

Faculty of Medicine and Health Sciences

Universiti Putra Malaysia

(Internal Examiner)

Zeehaida Mohamed, PhD

Lecturer

School of Medical and Health Sciences

Universiti Sains Malaysia

Malaysia

(External Examiner)

__________________________________

NORITAH OMAR, PhD

Associate Professor and Deputy Dean

School of Graduate Studies

Universiti Putra Malaysia

Date: 26 July 2011

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This thesis submitted to the Senate of Universiti Putra Malaysia and has been accepted as

fulfillment on the requirement for the degree of Master Sciences. The members of the

supervisory committee were as follows:

Rukman Awang Hamat

Lecturer

Faculty of Medicine and Health Sciences

Universiti Putra Malaysia

(Chairman)

Amal R. Nimir, PhD

Senior Lecturer

Faculty of Medicine

Cyberjaya University

(Member)

Ngah Zasmy A/L Unyah

Senior Lecturer

Faculty of Medicine and Health Sciences

Universiti Putra Malaysia

(Member)

__________________________________

HASANAH MOHD GHAZALI, PhD

Professor and Dean

School of Graduate Studies

Universiti Putra Malaysia

Date:

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DECLARATION

I declare that the thesis is my original work except for quotations and citations, which have been

duly acknowledged. I also declare that it has not been previously and is not concurrently,

submitted for any other degree at Universiti Putra Malaysia or at any other institutions.

__________________________________

LUBNA MOHAMED ELBESHTI

Date: 14 June 2011

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TABLE OF CONTENTS

Page

CHAPTER

1

2

GENERAL INTRODUCTION

1.1Objectives

1.1.1General Objectives

1.1.2 Specific Objectives

LITERATURE REVIEW

2.1 Taxonomy and Life cycle

2.1.1 Taxonomic Classification

2.1.2 Life Cycle

2.2 Medically Importance Cryptosporidium Species

in Human

2.3 Routs of Transmission

2.3.1 Waterborne Transmission

2.3.2 Food borne Transmission

2.3.3 Person to Person Transmission

2.3.4 Zoonosis and Other Routes of

Transmission

2.4 Immune Response and Pathophysiology of

Cryptosporidiosis

2.4.1 Immune Response to Cryptosporidium

2.4.2 Pathophysiology

2.5 Cryptosporidiosis in Immunocompetent

Children

2.5.1 Epidemiology

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ABSTRACT

ABSTRAK

ACKNOWLEDGEMENTS

APPROVAL

DECLARATION

LIST OF TABLES

LIST OF FIGURES

LIST OF ABBREVIATIONS

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2.5.2 Clinical Manifestations of

Cryptosporidiosis

2.5.3 Disease Outcomes

2.6 Cryptosporidiosis in Immunocompromised

Children

2.6.1 Epidemiology

2.6.2 Clinical Manifestations and Disease

Outcomes

2.7 Methods of Detection

2.7.1 Concentration Techniques

2.7.2 Staining Techniques

2.7.3 Immunological- Based Methods

2.7.4 Molecular Techniques

2.8 Therapy and Preventive Measures

2.8.1 Antimicrobial Therapy

2.8.2 Prevention and Control

METHODOLOGY

3.1 Study Site

3.2 Study Design

3.3 Study Duration

3.4 Study Population and Subject Selection

3.4.1Study Population

3.4.2 Sample Population

3.4.2.1 Inclusion Criteria

3.4.2.2 Exclusion Criteria

3.5 Sample Method

3.6 Sample Size

3.7 Data Collection

3.7.1 Questionnaire

3.7.2 Medical Record

3.7.3 Stool Samples Collection

3.7.4 Stool Samples Transportation

3.8 Samples Processing

3.8.1 Immunochromatographic (ICT) assays

RIDA-Quick ( R-Biopharm , Germany )

Procedure

3.8.2 Concentration Technique (Formalin-Ether)

3.8.3 Modified Ziehl- Neelsen Technique

3.8.4 Direct Fecal Smear Technique

3.8.5 Disposal of the Stool Samples

3.9 Study Ethics

3.10 Variables

3.10.1 Dependent Variables

3.10.2 Independent Variables

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3.11 Statistical Methods

3.12 Definition of Terms

3.12.1 Fever

3.12.2 Diarrhea

3.12.3 Anemia

3.12.4 Leucopenia

3.12.5 Neutropenia

3.12.6 Febrile Neutropenia

3.13.7 Risk Factors

3.13 Limitations

3.13.1 Difficult to get the approval from

Ministry of Health (MOH)

3.13.2 Missing of Some Questionnaires

3.13.3 Inadequacy of Some Medical Record

3.13.4 Difficult to Get Stool Samples From the

Patients

3.13.5 Insufficient the Volume of Some Stool

Samples

3.13.6 Time

3.14 Bias

3.13.1 Recall Bias

3.13.2 Information Bias

RESULTS

4.1 Detection of Intestinal Parasites in Stool

Samples Collected from Children with

Malignancies

4.2 Socio-demographic Characteristics of Children

with Malignancies

4.3 Clinical Characteristics and Laboratory of

Children with Malignancies

4.4 Type of Malignancies Diagnosed in Children

in Oncology Ward

4.5 Factors that Might Influence the Acquisition of

Cryptosporidiosis in Children with

Malignancies

DISCUSSION

CONCLUSION AND RECOMMENDATION

REFERENCES

APPENDICES

APPENDIX A: QUESTINNAIRE

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