The epidemiology of brucellosis in the Sultanate of Oman Abdulmajeed Al-Rawahi A thesis presented in fulfilment of the requirements for the degree of Doctor of Philosophy College of Veterinary Medicine School of Veterinary and Life Sciences Murdoch University Western Australia (August 2015) DECLARATION I declare that this thesis is my own account of my research and contains as its main content work which has not previously been submitted for a degree at any tertiary education institution Abdulmajeed Al-Rawahi ii ABSTRACT Brucellosis is a serious disease of cattle in many countries of the world, including Mediterranean and Middle Eastern countries. The study outlined in this thesis was conducted to investigate the epidemiology of brucellosis in the Sultanate of Oman. Thirty of 1267 holdings tested in the Sultanate contained seropositive animals for brucellosis (herd prevalence 2.4%, 95% CI 1.6, 3.4%). The southern governorate (Dhofar) had significantly more seropositive holdings (n = 20, 8.6%, 95% CI 5.3, 13) than did the northern governorates (n = 10, 0.97%, 95% CI 0.5, 1.8) (p < 0.001) highlighting the endemic nature of the disease in Dhofar. Although there were no significant differences between the herd seroprevalence for individual species, the highest herd level seroprevalence was reported in cattle (4.9%) followed by camels (2.3%), goats (1.4%) and sheep (0.6%). The overall individual animal seroprevalence of brucellosis in Oman was generally at a low level (0.4%; 95% CI 0.2, 0.5). The individual seroprevalence level in the different species was also low, being 0.4%, 0.4%, 0.4% and 0.1% in cattle, camels, goats and sheep, respectively. The practice of moving animals without testing between governorates is likely to have allowed the spread of infection throughout Oman. The active importation of live animals from other countries in the Horn of Africa, without prior monitoring of their brucellosis status, inter-species contact, sharing of common pasture, large herd size and the presence of poor biosecurity/unhygienic conditions in herds in the southern governorate may have facilitated the spread of brucellosis in the Dhofar region and from here infection may have been transmitted to other governorates. iii A logistic-regression analysis was undertaken to identify risk factors for disease. This analysis indicated associations of breed, age, herd size and production system with seropositivity. A higher seroprevalence was found in imported animals (OR 3.71, 95% CI 0.68, 20.43), and the seroprevalence increased with age. The latter finding is possibly because of a higher risk of contracting the disease after puberty through increased contacts with potentially infected animals. Only Brucella melitensis was cultured from different species of animals and biotype 1 was the only type identified in Oman by molecular means and phage typing. Sequencing of DNA revealed that all isolates had a very similar pattern. In the current study although there was no significant difference observed in the seroprevalence detected by different diagnostic assays (cELISA, iELISA and RBPT), the ELISAs were capable of detecting more positive samples than the RBPT and Rapid test. This may reflect the better sensitivity of the ELISAs and it is recommended that these tests be used in the control and eradication of brucellosis in Oman, where vaccination is undertaken. In Oman, human brucellosis was first reported in 1979 in the southern Dhofar governorate. A retrospective analysis of human brucellosis data sourced from the Ministry of Health, Oman from 1995 to 2012 was conducted. Information regarding location, age, gender, nationality of patients and year were included in the analysis. During this period, 2737 human cases of brucellosis were reported, with 96.7% of these in Dhofar. The incidence of disease was highest in young individuals (0-10 years of age), highlighting that these subjects were more at risk of acquiring brucellosis. The incidence of brucellosis was iv slightly higher in males (56%) than females (44%). Most of the positive patients were Omani nationals, most likely because of more opportunity for contact with infected animals on privately owned farms. The failure of disease control programmes in the southern region (2003 until 2012) could be due to a lack of information, inappropriate planning or administrative issues. With the information gathered from this study, it is considered there is a need to build a strategy to control the disease throughout Oman, rather than restricting control to the Dhofar governorate. However it is recommended that the control program adopted in the southern region (Dhofar), where the seroprevalence is high, be different to that implemented in the northern regions, where the disease prevalence is lower and more manageable. In the southern region, implementing a vaccination programme, along with individual animal identification and disease screening with a plan of intensive involvement and extension in the community, should be considered. In contrast in the northern region a test and slaughter program could be implemented. v ACKNOWLEDGMENTS I wish to express my deepest gratitude to my supervisor Professor Dr. Ian D Robertson for his guidance, encouragement, enthusiasm and constructive criticism throughout the development of this work. Deepest wishes and thanks extended to my co-advisor Dr. M. Saqib for his time and guidance. A special thanks are due to Dr. M. Hammad Hussain and his colleagues who supported me in all phases of this project, starting with the administrative and field work, the collection of samples, the statistical work and the use of GIS for disease mapping. Many thanks are also due to those technicians, veterinarians, drivers and workers who worked tirelessly under difficult conditions to assist me. I would like also to thank the Government of Oman, especially the Ministry of Agriculture and Fisheries, in providing the financial support for this study. And finally special appreciation must go to my family, relatives and friends in Oman and Australia who have encouraged and supported me over the years. vi TABLE OF CONTENTS DECLARATION ................................................................................................................. ii ABSTRACT ........................................................................................................................ iii ACKNOWLEDGMENTS ..................................................................................................vi TABLE OF CONTENTS .................................................................................................. vii LIST OF TABLES ..............................................................................................................ix LIST OF FIGURES .......................................................................................................... xii CHAPTER ONE .................................................................................................................. 1 INTRODUCTION ........................................................................................................ 1 1.1 Background ............................................................................................................. 1 1.2 Oman overview ....................................................................................................... 2 1.3 Issues ........................................................................................................................ 6 1.4 Objectives: ............................................................................................................. 13 CHAPTER TWO ............................................................................................................... 14 LITERATURE REVIEW .......................................................................................... 14 2.1 Etiologic Agent of Brucellosis .............................................................................. 14 2.2 History and Zoonotic Importance of Brucellosis ............................................... 16 2.3 Taxonomy of Brucellae ......................................................................................... 18 2.4 Clinical picture in animals and humans ............................................................. 27 2.5 Necropsy findings and microscopic lesions ........................................................ 33 2.6 Epidemiology of the disease ................................................................................. 34 2.7 Laboratory diagnosis ............................................................................................ 47 2.8 Treatment, prevention and control of Brucellosis ............................................. 65 CHAPTER THREE ........................................................................................................... 79 SEROPREVALENCE OF BRUCELLOSIS: CROSS-SECTIONAL STUDY ..... 79 3.1 Introduction ........................................................................................................... 79 3.2 Materials and Methods ......................................................................................... 79 3.3 Results .................................................................................................................. 102 3.4 Discussion ............................................................................................................ 129 vii CHAPTER FOUR ...........................................................................................................