Burkholderia Pseudomallei in Northern Vietnam
Total Page：16
File Type：pdf, Size：1020Kb
Burkholderia pseudomallei in Northern Vietnam: Environmental Detection and Molecular Characterization of Strains I n a u g u r a l d i s s e r t a t i o n zur Erlangung des akademischen Grades doctor rerum naturalium (Dr. rer. nat.) an der Mathematisch-Naturwissenschaftlichen Fakultät der Ernst-Moritz-Arndt-Universität Greifswald 2012 vorgelegt von Trinh, Thanh Trung geboren am 17.11.1980 in Ninh Binh, Vietnam Dekan: Prof. Dr. Klaus Fesser 1. Gutachter : Prof. Dr. Ivo Steinmetz 2. Gutachter : Prof. Dr. Leo Eberl Tag der Promotion: 12.09.2012 Contents Chapter 1 Background 4 Chapter 2 Clinical and microbiological features of melioidosis in 22 Northern Vietnam Chapter 3 Improved culture- based detection and quantification of 31 Burkholderia pseudomallei from soil Chapter 4 Highly sensitive detection and quantification of Burkholderia 38 pseudomallei in environmental soil samples using real-time PCR References 48 Declaration 57 Curriculum Vitae, Publications, Contributions to articles 58 Acknowledgements 61 Appendix 63 Chapter 1 Chapter 1 Background Trinh Thanh Trung 1. Introduction ........................................................................................................................ 5 1.1. Melioidosis ..................................................................................................................... 5 1.1.1. From origins to global distribution ......................................................................... 5 1.1.2. Clinical features of melioidosis .............................................................................. 6 1.1.3. Diagnosis and management of melioidosis ............................................................ 7 1.2. Characteristics of Burkholderia pseudomallei ............................................................... 9 1.2.1. Taxonomy of Burkholderia pseudomallei and closely related species .................. 9 1.2.2. Modes of acquisition and bioweapon potential .................................................... 11 1.2.3. Bacterial genome and its plasticity ....................................................................... 12 1.2.4. Bacterial genotype and population structure ........................................................ 15 1.3. B. pseudomallei in the environment ............................................................................. 17 1.3.1. Natural ecological niches and bacterial survival .................................................. 17 1.3.2. Methods for detection of B. pseudomallei from the environment ........................ 19 1.4. What do we know about melioidosis in Vietnam? ....................................................... 20 2. Aims of the study .............................................................................................................. 21 4 Chapter 1 1. Introduction 1.1. Melioidosis 1.1.1. From origins to global distribution Melioidosis, a term derived from the Greek “ melis ” (meaning donkey distemper) and “ eidos ” (meaning resemblance), is a serious infectious disease caused by a soil- dwelling Gram-negative bacterium Burkholderia pseudomallei (White, 2003; Cheng and Currie, 2005). A century ago, the first cases of the disease were described by the pathologist Alfred Whitmore in Myanmar (White, 2003). Several years later, a large number of human and animal melioidosis cases had been also detected Myanmar and Malaysia, respectively (Dance, 1991; White, 2003). During and after World War II, many sporadic cases of disease were recorded among the French and American soldiers who were stationed in Southeast Asia (Dance, 1991; Cheng and Currie, 2005). Subsequently, a large number of cases, including many epidemiologically linked clusters, were reported for indigenous populations in Thailand and Australia, where the first cases had been recognized in 1949 and 1956, respectively (Cheng and Currie, 2005). By the second half of the 20 th century, melioidosis had emerged as an important public health problem in certain areas of the tropics. In Ubon Ratchathani, Thailand, melioidosis accounts for up to 20% of community-acquired septicemic infection (Chaowagul et al., 1989) with an average annual incidence of 4.4 cases per 100,000 population. The mortality rate reaches approximately 40% in septic shock cases (Suputtamongkol et al., 1999). A study in Northern Australia also revealed a considerable percentage of melioidosis in community-acquired septicemic patients admitted to the Royal Darwin Hospital (Douglas et al., 2004). The average annual incidence of melioidosis in this region is 19.6 cases per 100,000 population and the overall mortality rate amounts to 16.2% among the infected cases (Currie et al., 2004). During the last 20 years, many attempts have been made to increase the awareness of melioidosis in routine diagnostic settings. Not only in Southeast Asia and Northern Australia, but also in different geographical areas of the world, many more cases of infection have been detected, resulting in the expansion of the global map of the disease (Figure 1-1). Sporadic and outbreak cases of melioidosis in Southern Taiwan as well as mainland China have been described (Yang, 2000; Shih et al. , 2009). Multiple cases have been reported from different regions of the Indian subcontinent (John et al., 1996). The occurrence of melioidosis patients as well as the presence of B. pseudomallei in the environment have been noted in Brazil (Rolim et al., 2005). 5 Chapter 1 Figure 1-1. Worldwide distribution of melioidosis (from Currie et al., 2008) 1.1.2. Clinical features of melioidosis Melioidosis has a wide variety of clinical presentations, ranging from an asymptomatic carrier state to a chronic debilitating localized infection or acute fulminant septicemia (White, 2003). Multiple organs or even only single sites of the body may be involved during the febrile episodes of infection (Leelarasamee, 2004). Therefore, clinical signs and symptoms at the onset of the disease vary, depending on the affected organs (White, 2003), leading to unpredictable clinical manifestations and resulting in the misdiagnosis of melioidosis as other infectious diseases, such as tuberculosis or typhoid fever in clinical settings (Stone, 2007; Vidyalakshmi et al. , 2008). In all of the hospitalized cases, septicemia is the most common presentation of melioidosis, and the most commonly affected organ is the lung (White, 2003). Skin and soft-tissue abscesses are also common sites and may be the source of systemic infection or the result of hematogenous spread (Cheng and Currie, 2005). In a few cases, melioidosis presents as uncomplicated localized infections of the skin, subcutaneous tissues, or eyes (White, 2003; Gal et al. , 2004). Additionally, abscess formation can occur in any internal organ, such as liver, spleen, lymph nodes, prostate, and skeletal muscles. In renal abscesses, melioidosis is often associated with genitourinary infection (White, 2003). Moreover, clinical presentations of endocarditis, peritonitis, osteomyelitis or septic arthritis, encephalitis or meningitis, and 6 Chapter 1 mycotic aneurysm have been recently seen in many melioidosis patients (Hsueh et al., 2001; Malczewski et al., 2005; Vidyalakshmi et al., 2008). A number of common risk factors for developing melioidosis have been defined. They include diabetes mellitus, chronic renal disease, chronic lung disease, excess alcohol uptake, and thalassemia (Suputtamongkol et al., 1999; Currie et al., 2004). Of thes, diabetes mellitus has the highest incidence rate for melioidosis; the estimated rate for meliodosis infection is 260.4 cases per 100,000 diabetic patients (Currie et al., 2004) and the underlying diabetes disease in melioidosis patients ranges from 37 to 64% (Malczewski et al., 2005). Moreover, other independent risk factors, such as malignant tumors and steroid therapy for immunosuppression, are also reported for melioidosis (Suputtamongkol et al. , 1999). Age above 45 years and occupation of farmer or construction worker – who are directly exposed to contaminated soil and water – are predisposing factors for developing melioidosis (Suputtamongkol et al., 1999; Currie et al., 2004). Recently, tsunami natural disaster in December 2004 has been associated with large numbers of detected cases, suggesting tsunamis as a new risk factor for melioidosis (Peacock, 2006). In different regions, the overall mortality rates of melioidosis are approximately 20 to 50%. Death occurs within 48 h after admission in most of the cases with disseminated bacteremia and septic shock (Simpson et al., 1999; Currie et al., 2000; Limmathurotsakul et al., 2006). Recurrent melioidosis has been reported in 13.1% of recovered patients. Of these, 75% develop the disease by relapse. The others manifest the disease by reinfection (Limmathurotsakul et al., 2006). Besides humans, melioidosis can affect a wide range of animals. Sporadic and outbreak cases have been reported in pets, such as dogs and cats, in farm animals, such as horses, mules and donkeys, goats, sheep, cattle, and buffalo, in zoo animals such as seals, dolphins, crocodiles, and also in human primates and non-human primates (Sprague and Neubauer, 2004). 1.1.3. Diagnosis and management of melioidosis Because of a wide range of clinical presentations, diagnostic confirmation of a melioidosis case has to depend heavily on clinical microbiological examination (Inglis et al., 2006). Although many antigen or antibody detection assays and molecular techniques have been developed for diagnosis of melioidosis, culture-based