Sero-Survey of West Nile Virus and Japanese Encephalitis Virus in Two Provinces of Pakistan

摘要

This study describes the first large-scale sero-survey on West Nile virus (WNV) conducted in the equine population in Pakistan.Sera were collected from 449 equids from two provinces of Pakistan during 2012-2013.Four prefectures from Punjab province and two from KPP province were selected on the density of the draught equids, migratory birds, presence of wetlands and veterinary clinics Equine serum samples were screened using a commercial ELISA kit detecting antibodies against WNV and related flaviviruses.ELISA-positive samples were further investigated using virus-specific microneutralization test (MNT) to identify infections with Japanese encephalitis virus (JEV), WNV and tick-borne encephalitis virus (TBEV).
　　Anti-WNV antibodies were detected in 292 samples by ELISA (seroprevalence 65.0％) and WNV infections were confirmed in 249 animals by MNT.However, there was no animal found infected by JEV or TBEV.Significantly (P ＜ 0.001) higher prevalence was observed in equines from Punjab (79.4％) compared to those from KPK (49.3％).In Punjab province higher seroprevalence of antibodies was observed in equines of Lahore (84.6％), followed by Sargodha (83.3％), Gujranwala (81.2％) and Faisalabad (71.9％).In KPK province significantly higher (P ＜ 0.001) prevalence was observed in equine from Peshawar (62.8％) compared to Charsadda (35.1％).Prevalence of WNV antibodies was not found significantly different (P =0.216) between two sampled species: i.e., donkeys (51.2％) and horses (58.9％).Age was found to be significantly (P＜0.05) associated with the seropositivity of WNV in equines.Seroprevalence recorded in young ＜ 5 years), adult ＜ 10 years) and old (＞ 10 years) equines was 72％, 59.3％ and 50％ respectively.Out of WNV positive animals 153 (56.4％) were females and 96 were males (53.9％).Prevalence of WNV was not found significantly different between male and female equids.The detection of WNV-seropositive equines in Pakistan strongly suggests a widespread circulation of WNV in Pakistan.

目录

声明

CONTENTS

ABSTRACT

Chapter 1 INTRODUCTION

1．1．EVOLUTION OF FLAVIVIRUS

1．2．FLAVIVIRUS GENOME AND PROTEINS

1．2．1 Flaviviral RNA synthesis and replication

1．2．2 Structural Proteins

1．2．3．Non Structural Proteins(NS)

1．3．JAPANESE ENCEPHALITIS SEROLoGICAL GROUP OF FLAVIVIRUS

1．4．JAPANESE ENCEPHALITIS VIRUS(JEV)

1．4．1．Transmission of JEV

1．4．2．Pathogenesis of JEV

1．4．3．Epidemiology of JEV

1．5．WEST NILE VIRUS(WNV)

1．5．1．Transmission of WNV

1．5．2．Pathogenesis of WNV

1．5．3．Global Epidemiology of WNV

1．6．VIRUS DETECTION

1．6．1．Surveillance for WNV and Other Arboviruses

1．7．SERODIAGNOSIS OF WNV ANDRELATED FLAVIVIRUS IN EQUINE

1．8．AIMS OF THIS THESIS

Chapter 2 MATERIALS AND METHODS

2．1 STUDY LOCALES AND SETTINGS

2．2 SAMPLING FRAME

2．3 SELECTION OF ANIMALS

2．4 COLLECTION ANDTR ANSPORTATION OF BLOOD SAMPLES

2．5 SEPARATION ANDPRESERVATION OF SERUM

2．6 SERO-DIAGNOSIS OF WNV ANDJEV

2．7 DESCRIPTION OF THE TEST

2．8 DESCRIPTION AND PRINCIPLE

2．8．1 Sample Preparation

2．8．2 Wash Solution Preparation

2．8．3 Testing Procedure

2．8．4 Validation

2．8．5 Interpretation

2．9 Microneutralization Test(MNT)

2．9．1 Materials and Equipment

2．9．2 Chemicals and Reagents

2．9．3 Testing Procedure

2．9．4 Interpretation

2．10 Statistical analysis

Chapter 2 RESULTS

3．1．GEOGRAPHIC ANDCLIMATIC DESCRIPTION OF SELECTED AREAS

3．1．1．Punjab

3．1．2．Khyber Pakhtunkhwa

3．2．EQUINE POPULATION IN STUDY AREAS

3．3．DEMOGRAPHIC CHARACTERISTICS OF THE SAMPLED POPULATION

3．3．1．Spatial distribution of equines(n=449)for sero-prevalence of WNV and JEV in selected districts

3．3．2．Equine type related distribution of equines(n=449)for sero-prevalence of WNV and JEV in selected areas

3．3．3．Sex related distribution of samples(n=449)for sero-prevalence of WNVand JEV in selected areas

3．3．4．Age related distribution of samples(n=449)for sero-prevalence of WNV and JEV in selected areas

3．4．ELISA BASED SEROLOGICAL PREVALENCE OF WEST NILE VIRUS AND RELATEDFLAVIVIRUSES

3．4．1．Serological prevalence of West Nile virus and related flaviviruses by ELISA

3．4．2．Area related Seroprevalence

3．4．3．Seroprevalence of antibodies in two different species of equids

3．4．4．Age related seroprevalenee of viral antibodies

3．4．5．Sex related seroprevalence of viral antibodies

3．5．MNT BASED SEROLOGICAL PREVALENCE OF WEST NILE VIRUS，JAPANESE ENCEPHALITIS VIRUS ANDTICK BORNE ENCEPHALITIS VIRUS

3．5．1．Serological prevalence of West Nile virus and related flaviviruses by MNT

3．5．2．Area related seroprevalence of West Nile virus

3．5．3．Seroprevalence of West Nile virus specific antibodies in two different species of equids

3．5．4．Age related seroprevalence of WNV specific antibodies

3．5．5．Sex related seroprevalence of anti-West Nile virus antibodies

Chapter 4 DISCUSSION

Chapter 5 CONCLUSION

LITERATURE CITED

PAPERS PUBLISHED

ACKNOWLEDGEMENT