Melioidosis – A Serious Emerging Threat in Bangladesh

Melioidosis, a pyogenic infection that presents acutely or as a chronic infection, is caused by the soilassociated bacterium Burkholderiapseudomallei. Infection is acquired by inoculation or inhalation and is more common in patients with underlying chronic disease. It is endemic in the tropical belt. Although Bangladesh is not considered as a country where melioidosis is endemic, an increasing number of cases have been reported recently. Definitive diagnosis requires the isolation of B. pseudomalleiin culture from clinical specimens. However, the laboratory diagnosis of melioidosis in Bangladesh and other under-resourced countries is limited by a lack of familiarity with the bacterium and a lack of facilities to accurately confirm the identity of the isolate. It is highly likely that melioidosis is underdiagnosed in this country. There is a need to increase awareness of this infection among clinicians and clinical microbiologists and improve laboratory facilities for the selective isolation and accurate identification of B. pseudomallei. Melioidosis has a notoriously protracted course; cure is difficult without a prolonged course of appropriate antibiotics.


Background
Melioidosisis a collective term for infection caused by the soil organism Burkholderiapseudomallei.The causative organism was first described by Whitmore in 1912 when hefirst isolated B. pseudomalleifrom an opiate addict in Rangoon. 1 Whitmore's name was for some time eponymously linked with the disease melioidosis.So,name meloidosis, also known as Whitemore disease, is taken from 'melis' meaning 'distemper of asses' and 'eidos' meaning resembles 'glanders' 2 .For many years the causative organism of melioidosis was classified within the Pseudomonasgenus; however, in 1992,along with P. mallei and four other species, P. pseudomalleiwas reclassified to a new genus named after the USmicrobiologist Walter Burkholder.The genus Burkholderiacomprises at least 12 species, many of which are natural inhabitants of the rhizosphere, the bacteriological and chemical milieu of plant roots 3 .B. pseudomalleihas been classified by the Centers for Disease Control and Prevention as a category B bioterrorism agent, resulting in increased research and understanding of melioidosis 4 .The first case of human melioidosis in Bangladesh was described in a diabetic adult in 2001.Since 2001, 5 cases of meloidosis were diagnosed in Ibrahim Medical College.All are diabetic and hailing from Mynensing,Tangail and Gazipur.Recently, scientist found this organism in the soil of gazipurfor the first time in Bangladesh 5 .Therefore, Melioidosis is an uncommon but maybe fatal tropicaland emerging infectious diseasein Bangladesh.Its true prevalence however is not known, as there is under-reporting of its incidence due to the poorly understood disease process and misdiagnosis.At the same time enough resources are not always available in some areas to carry out research and increase the awareness of the general public and to educate and familiarize the medical profession about the disease.So, by this article, we try to increase the awareness about this disease among physicians.

What is its Spectrum?
The geographic area of the prevalence of the organism is bound to increase as the awareness increases.This disease has emerged over the past 25 years as an important cause of morbidity and mortality in Southeast Asia and northern Australia, and is also endemic in other tropical regions 6 .Melioidosis occurs predominantly in Southeast Asia, northern Australia, South Asia (including India), and China 7- 12 .The majority of diagnosed cases are from Thailand 13- 16 .Malaysia [17][18][19][20] ,Singapore [21][22][23][24][25][26][27] and northern Australia 28- 30 .Cases are also reported from Papua New Guinea 31 and New Caledonia 32 .Northeastern Thailand and parts of northern Australia are "hyperendemic" for melioidosis 14,28 with seasonal peaks in the wet seasons.In 2010, there was been an increase in incidence in both northeast Thailand and northern Australia as well as in south Asia [33][34][35] In Thailand 2000 to 3000 new cases are diagnosed every year. 36n Malaysia, reported seroprevalence in healthy individuals is 17-22% in farmers ( mainly rice farmers)and 26% in blood donors. 37In North Australia 0.6 to 16% of children have evidence of infection by B. pseudomallei . 28lioidosis has been described outside the classic endemic regions.Most of such cases are acquired by visitors to endemic areas, with symptoms arising later following departure from the endemic area.However, sporadic human or animal cases and occasional environmental isolates of B. pseudomallei have been described from Africa, Indian Ocean countries (such as Mauritius), the Middle East, the Caribbean, and Central and South America [38][39][40] Some of these reports represent incorrect species diagnosis, but others have been confirmed as B. pseudomallei, making the endemic boundaries of melioidosis less clear [10][11][12] .
The B. pseudomallei have also been isolated in America.There are reports ofseveral cases of patients with melioidosis who have immigrated into Europe and the disease has been increasingly recognized in returning travellers to Europe from endemic areas. 41Two cases of melioidosis were reported in 2005 from southern Florida; both patients likely had separate exposure in Honduras . 42[45][46] Melioidosis is an emerging infection in India, with a reported prevalence of 7%. 47Sri Lanka, positioned between 5-10 0 N, is situated in the endemic belt and has similarities in weather and environmental conditions with these countries.However Sri Lanka has been considered non-endemic for melioidosis. 48lioidosis affects all ages but peak incidence is mainly between 40 to 60 years of age, with male to female ratio of 1.4:1. 49There is a good correlation between the isolation of the organism from soil and the seroprevalence of antibodies in the population living in that region.

What is B. pseudomallei ?
B. pseudomallei is agram negative intracellular organism, natural inhabitant of soil and water in the tropics and subtropics but can also survive in dry atmospheric conditions.It is ubiquitous in the rice-farming areas.It is also present in rubber plantations, cleared fields, cultivated and irrigated agricultural sites as well as drains and ditches.When isolated from blood, sputum, pus and other body fluids, B. pseudomalleiappears like safety pins (bipolar) under the microscope with methylene blue stain.It grows aerobically on ordinary media at 37 0 C. Colonies are wrinkled and show dry daisy-head appearance along with a distinct odour.Mucoid colonies suggest that the patient is receiving antibiotic therapy.
B. pseudomallei can survive anaerobic conditions in the presence of acidic environment, and also survive in distilled water for several years. 50.The bacterium is resistant to penicillin, aminoglycosides, rifamycins and relatively insensitive to quinolones and macrolides.Therefore the therapeutic options are limited and continuous presence of the organism in patients is not fully understood. 51,52B. pseudomalleiis resistant to macrolide and aminoglycoside antibiotics via a multidrug efflux pump.Mutations within the conserved motifs of the beta-lactamase enzyme ( enzyme that hydrolyses the cyclic amide bond of beta-lactam antibiotics )also account for the resistance patterns. 53

How does it transmit?
There are several established modes of transmission within the patient population.The possible modes are inhalation, ingestion or inoculation through the skin lesions from the contaminated soil. 54Person-to-person transmission of B. pseudomallei especially between patient and his sibling or one of their playmates is common. 55Vertical transmission(from mother to child) is possible. 56It can also be transmitted by direct contact with infected rodents or infected food, soil, water, excreta; person-to-person transmission is also possible through use of injection needle.B. pseudomallei can also be transmitted through sexual intercourse. 57The link between melioidosis and consumption of Kava (Piper methysticum) has also been seen. 57Heavy rains and winds may cause increased inhalation of B. pseudomallei .Interestingly, a container of commercial handwash detergent was a source of infection in Northern Australia. 29w does the disease occur?B. pseudomallei attack several eukaryotic cell lines..60Capsule and a type III secretion system (TTSSexpressed mainly by pathogenic bacteria that is used to introduce deleterious proteins called effectors into host cells )facilitate B. pseudomallei to survive, escape from endocytic vesicles, facilitate bacterial invasion of epithelial cells and intracellular survival. 61,62The uptake of B. pseudomallei by several cell lines in culture leads to induction of cell fusion and formation of a multinucleated large cell. 65Production of nitric oxide has bactericidal activity and failure of infected cells to successfully control the growth and subsequent survival of intracellular B. pseudomallei are due to the suppression of inducible nitric oxide synthase ( iNOS ) by B. pseudomallei . 63,64However, interferons enhance antimicrobial activity of macrophage infected B. pseudomallei by up-regulating iNOS. 65,o are at risk? Diabetis mellitus, Excessive alcohol consumption, Chronic renal impairment, Cystic fibrosis, Chronic heart failure, Chronic pulmonary disease,Leukaemia, lymphoma, Corticosteroid therapy, Immunodeficiency, Neoplasm and Kava Consumption 12 .

What are the clinical manifestations?
It is important to note that melioidosis has a wide range of signs and symptoms that can be mistaken for other diseases such as tuberculosis or more common forms of pneumonia.Clinical manifestations of melioidosis range from localised infection to acute pneumonia and fulminant septic melioidosis. 66,67,68B. pseudomallei can cause disease in apparently healthy individuals .Once infected, it may remain dormant and become active after months, years or decades when host is immunocompromised.The factors that provoke the reactivation of latent pathogen probably are environmental variables, stress and immunity status. 69,70ocalized melioidosis occurs in the form of acute suppurative lesions, superficial and deep-seated abscess in the psoas muscle, parotid glands and at the root of mesentery. 71,72It may also present as cellulitis, chronic otitis media and sepsis after burns and trauma. 73,74,The other manifestations are mycoticaneurysm, pericarditis, osteomyelitis epididymoorchitis and prostatitis. 75,76,77Melioidosis is also associated with systemic lupus erythematosus. 78Melioidotic prostatic abscesses are reported very rarely and are not easy to diagnose.In endemic areas, the elderly diabetic person who presents with high-grade fever and urinary obstruction may have B. pseudomalleiin the prostate gland 79 .Central nervous system involvement including brain abscess is a rare complication with high mortality. 80,81The immunesuppressed patients present with melioidosis septicaemia and their clinical features are similar to other gram-negative septicaemias and its prognosis is poor.

How does it investigated?
The diagnosis of acute or chronic melioidosis remains challenging.In endemic areas, melioidosis should be considered in the differential diagnosis of any Pyrexia of Unknown Origin (PUO), acute respiratory distress syndrome (ARDS) and acute septicaemia.The other conditions that melioidosis may present as are pneumonia, acute suppurative lesions, chronic granulomatous lesions, septic arthritis, osteomyelitis, epididymorchitis and mycotic aneurysm as well as radiological pattern of tuberculosis on the chest X-ray but not supplemented with mycobacterium tuberculosis positive sputum culture.In melioidosis, laboratory diagnosis is essential for successful patient management.C-reactive protein (CRP), an early indicator of infectious or inflammatory conditions may be elevated in melioidosis; however under normal CRP levels, melioidosis should not be ruled out. 82

Identification of B. pseudomallei
Isolation of B. pseudomalleiby culture from a clinical specimen [blood, urine, sputum, skin lesions and swab samples from throat] is the gold standard of diagnosis. 83 B. pseudomallei that appears in the blood within 1-2 weeks after the infection and reach maximal titre in 4 to 5 months. 88However its interpretation may be difficult because of the following points; • False positive results due to cross-reaction with other gram negative bacteria which shares antigens (lipopolysaccharide of cell wall) particularly Burkholderiacepeciaand Legionella species.
• There may be rare false negative results • High antibody titre may persist for a long time after infection subsides.
Enzyme linked immunosorbent assay (ELISA)test detects specific IgG and IgM antibodies of B. pseudomallei in serum specimens.ELISA is more convincing in terms of sensitivity and specificity for antibody detection as it points to an active disease process. 89The indirect ELISA is easy to perform and hence is recommended as a diagnostic serological test when melioidosis is in the differential diagnosis of PUO cases.Immunoflurorescent Antibody Assay is a rapid, highly sensitive and specific test for the identification of current infection.

Molecular identification techniques
Molecular biology techniques such as polymerase chain reaction (PCR), dot immunoassay, pulsed field gel electrophoresis (PFGE), restricted fragmentation length polymorphism (RFLP) and random amplification of particle of deoxyribonulease (RAPD) are also used for diagnosis.These are the recommended techniques for the rapid diagnosis of the disease and for monitoring therapy and epidemiological studies because of its high sensitivity, specificity, simplicity and speed.In recent times sensitive PCR amplification techniques for detecting the DNA of B. pseudomallei in clinical specimens, especially buffy coat specimens of acute melioidosis patients have been useful. 90,91boratory diagnostic approach How it is treated?
The main objective of treatment is to reduce the mortality and morbidity in melioidosis.Before the advent of proper antimicrobials, the mortality of the melioidosis patients used to be around 95%. Rational use of antimicrobials has reduced it to half.B. pseudomalleiis inherently resistant to penicillin, ampicillin, first-generation and second-generation cephalosporins, gentamicin, tobramycin, streptomycin, and polymyxin.Of the newer antibiotics, ertapenem, tigecycline, and moxifloxacin have limited in vitro activity against clinical isolates of B. pseudomallei, and the minimum inhibitory concentration for doripenem is similar to that for meropenem. 92Various mechanisms of acquired antibiotic resistance have been identified, including efflux pumps, enzymatic inactivation, bacterial-cell-membrane impermeability, alterations in the antibiotic target site, and amino acid changes in penA, the gene encoding the highly conserved class A â-lactamase. 93,94eatment is divided into intravenous and oral phases.Initial parenteral therapy is given for 10-14days or until clinical response is seen (whichever is the longer).Ceftazidime or a carbapenem antibiotic is the treatment of choice.Ceftazidime is used as first-line therapy in Thailand, with a switch to acarbapenem antibiotic in the event of treatment failure on ceftazidime.Parenteral treatment at the Royal Darwin Hospital, Australia (which sees the highest number of cases in Australia) consists of ceftazidime,or meropenem plus G-CSF if the patient has septic shock. 95e use of G-CSF in patients with severe melioidosis in Thailand is not supported by published evidence. 96The results of an ongoing randomized trial of ceftazidime versus meropenem for the treatment of melioidosis in Thailand will not be available for several years.The routine addition of TMP-SMX to ceftazidime or meropenem during the initial intensive therapy phase was discontinued in 2005. 97TMP-SMX is usually used in Australia for patients with neurological or prostatic melioidosis in view of its excellent penetration, the evidence for which is based on expert opinion and case series. 98travenous amoxicillin-clavulanic acid (AMC) is second-line empiric treatment.The switch from parenteral to oral antimicrobial therapy is made once the patient shows clear evidence of clinical improvement, including an absence of fever for 48 h and negative repeat blood culture taken at around 1 week after the onset of therapy.Prolonged parenteral therapy may be required for patients with disseminated infection, involvement of the central nervous system, bone or joint, and patients with deep-seated abscesses that cannot be drained.
Oral therapy consists of TMP-SMX alone (Australia) or in combination with doxycycline (adults in Thailand).Results are pending of arandomized controlled trial, which has recently been completed in Thailand to determine whether TMP-SMX and TMP-SMX plus doxycycline are equivalent.
AMC is an alternative for patients with intolerance to TMP-SMX and is first-line therapy for children and pregnant women in Thailand, but is associated with an increased risk of relapse compared with TMP-SMX-based therapies. 33armacodynamic and pharmacokinetic modelling indicate that the recommended AMC dose should be 20/5 mg/kg every 8 h. 99Twice daily doses or formulations containing AMC ratios .4 to 1 are notrecommended. 100Chloramphenicol is no longer recommended for the treatment of melioidosis. 101Its use in current clinical practice is extremely rare and reserved for neurological infection if ceftazidime, carbapenems or trimethoprim-sulphamethoxazole cannot be used. 102In resource-poor settings where parenteral therapy is often difficult to provide or sustain, patients may be treated with oral antimicrobialdrugs.Under such circumstances, the regimen prescribed will be dictated by drug availability and cost, and chloramphenicol may form a component of treatment.
The recommended duration of oral treatment is 3-6 months.For patients with hepatosplenic abscesses, duration of therapy should be guided by time to resolution on serial abdominal imaging.It is not known whether a shorter course of therapy may be adequate for patients with mild and localized disease, such as a single subcutaneous abscess.Monitoring of drug adherence is crucial, as this is probablythe most important factor in determining recurrence.

Treatment recommendations
How long treatment is required?Appropriate treatment is imperative in order to prevent relapse and failure of treatment.Despite appropriate treatment, melioidosis has a higher relapse rate.The average time between discharge from hospital and relapse is of 21 weeks.Treated patients require long-term follow up, as B. pseudomalleiremains latent for up to 26 years in the body. 103For maintenance therapy, Co-Amoxyclav is a safe and well-tolerated antimicrobial agent (there is some concern that it may be less effective than theconventional regimen of chloramphenicol, co-trimoxazole and doxycycline).The recommended duration for maintenance therapy is of 12 to 20 weeks.pseudomallei it is important that milk is pasteurized before consumption. 108ere is currently no licensed vaccine available for protection against melioidosis.At present studies are underway to identify possible antigens using lipopolysacchrides of B. pseudomallei in mouse models. 109Antibodies against B. pseudomalleiflagellin reduce the motility of the bacterium and provide protection against melioidosis in animal models. 110A recent study has shown that quicklime was able to inhibit the growth of B. pseudomallei in soil from a rice field. 111As our understanding of the disease increases and as we move forward with the studies on the pathogenesis of the disease, new and effective vaccine against melioidosis may become a reality.

Conclusion:
A high index of suspicion is required in order to diagnose melioidosisin the non-endemic setting.Clinicians should consider the possibility inpatients with a fever who have one or more of the following: a historyof residency in, or travel to a region where melioidosis is endemic; anoccupation or other pursuits associated with contact with soil or waterthat might contain B. pseudomallei(including military personnel whoare on exercise or active service); and the presence of risk factors suchas diabetes mellitus or renal disease.The variability in clinical featuresof infection is such that it is often impossible on clinical grounds todifferentiate between melioidosis and other acute and chronic bacterial infectionsspecially tuberculosis.Confirmation of the diagnosis relies on good practicesfor specimen collection, laboratory investigations.
84rrect identification of B. pseudomalleiis essential for long term supportive therapy in the treatment of melioidosis.A few simple tests can be employed to identify B. pseudomalleiin the endemic areas.These tests include positive oxidase test, bipolar gram staining, metallic sheen colonies on special media (Ashdown media which contains various dyes and gentamicin) and resistance to aminoglycosides.84 Skin and soft tissueCellulitis, Subcutaneous abscess,Infected wound, Chronic granuloma, Ecthyemaagangrenosum, Haemorrhageic bleb, Chronic pustules, Pyomiositis, urticaria, mastitis.
infection through the feet and hands.It is important to maintain safe water through regular disinfection and safe storage of water for both human and animals bred for human consumption.Sewage wastes can attract insects and rodents and encourage the growth of B. pseudomallei.Therefore proper disposal of sewage wastes is essential in endemic areas.As dairy products can contain B.