Chapter 37 Melioidosis in Marine Mammals
Melioidosis is the disease caused by Burkholderia pseudomallei (previously Pseudomonas pseudomallei), a gram-negative bacillus. The closest relative to B. pseudomallei is B. mallei, the etiologic agent of glanders in horses, and a zoonosis. Unlike B. mallei, however, B. pseudomallei is ubiquitous in soil and water in endemic areas (of Southeast Asia and Northern Australia). Burkholderia pseudomallei was first identified in Burma in 1911 in a 10-year-old male morphine addict with lung abscesses.60 It has subsequently become an important cause of community-acquired disease in humans in some endemic regions.10,14
Melioidosis affects a wide range of animal species, including laboratory animals, livestock (pigs, sheep, goats, cattle), companion animals (horses, dogs, cats), zoo and wildlife animals (marsupials, camelids, primates, deer, birds), and even perhaps cold-blooded animals (crocodiles, snakes, tortoises).6,12,35,37,55 However, species susceptibilities to and manifestations of infection differ. Marine mammals appear to be especially susceptible; five species of cetaceans (Tursiops gilli, Tursiops aduncus, Orcinus orca, Pseudorca crassidens, and Lagenorhynchus obliquidens) and eight species of pinnipeds (Zalophus californianus, Eumetopias jubatus, Arctocephalus pusillus, Neophoca cinerea, Phoca vitulina, Mirounga leonine, Hydrurga leptonyx, and Halichoerus grypus) have been reported to have melioidosis.13,33 Almost all reported cases of melioidosis in marine mammals are from Ocean Park, an oceanarium in Hong Kong.
Burkholderia pseudomallei is an aerobic, non–spore-forming, gram-negative bacillus that appears as short rods with bipolar staining, giving it a “safety pin” appearance. It has two or more polar flagellae and is motile. The organism grows on standard laboratory media such as blood agar and, more selectively, on MacConkey agar.22 However, the selective medium of choice is Ashdown medium, which contains crystal violet and gentamicin.4,63 The morphology of colonies of B. pseudomallei may be smooth and mucoid or, more distinctively, dry and wrinkled after incubation for a few days. Colonies have a purple color if grown on Ashdown medium. The organism also has a pungent “earthy” odor and forms a pedicle when grown in broth. Biochemical tests used to identify B. pseudomallei include a positive oxidase reaction, production of gas from nitrate, activity of arginine dihydrolase plus gelatinase, and the oxidation of a variety of carbohydrates.22 The Rapid NFT commercial kit by BioMerieux, API 20NE, has been evaluated to be greater than 99% accurate in identifying B. pseudomallei.18 Burkholderia pseudomallei is a containment level 3 organism.1
EPIDEMIOLOGY AND EPIZOOTIOLOGY
Southeast Asia and Northern (tropical) Australia are the major endemic foci for melioidosis. Endemic areas generally lie between the latitudes of 20°N and 20°S and have a tropical or subtropical climate.26 Burkholderia pseudomallei is an environmental saprophyte, surviving in soil and water, and often may be isolated from the environment in endemic areas, although the distribution of the organism is uneven. Climate (rainfall, temperature, sunlight); soil composition, type, and chemistry; and the interaction of B. pseudomallei with other biologic organisms (e.g., plants, other soil microflora) may affect the multiplication and survival of B. pseudomallei in its environment, thereby determining its distribution and the prevalence of disease. Moisture and temperature are two significant environmental factors. Endemic areas have high rainfall and temperatures, and both environmental B. pseudomallei and melioidosis are rare in countries with low rainfall.17
Infection is primarily through exposure to soil or water contaminated with B. pseudomallei.17,36 In humans the methods of natural infection are inoculation, inhalation, and possibly ingestion, with inoculation considered the most common. However, the mode of infection could be documented in only 5.7% of cases in Thailand.49 The primary route of infection in marine mammals remains uncertain. The incubation period for melioidosis is also not known in marine mammals. Reports in humans range from 2 to 3 days20 to “a few days to weeks.”15 It is believed that the acute fulminating septicemic form of melioidosis has a short incubation period and short course.59
Melioidosis was first described in Hong Kong in an outbreak in dolphins at Ocean Park in 1975, resulting in the sudden death of 24 dolphins.27 Human melioidosis started being recognized in Hong Kong after this first report,45,61 although the prevalence of disease is apparently low, with an estimated 10 persons/year diagnosed in the territory.25 Available records show that at least 78 marine mammals have died from melioidosis at Ocean Park since it started maintaining these animals in 1974. This number is probably underestimated because it does not even include the 24 dolphins cited in Huang’s paper.27 Only cases that could be definitively diagnosed (i.e., positive culture of B. pseudomallei) were included in epidemiologic studies, and records for these dolphins were limited. These mortalities reflect the scale, importance, and significance of melioidosis at this facility. Epidemiologic investigations continue to be conducted at Ocean Park33 to try to understand the disease better and formulate preventive measures. Findings include the following:
Increased prevalence of disease during the wet monsoon season in endemic regions is also a typical finding in human cases.14,29,49 Cases in animals have also been associated with flooding in Australia.30,31 To determine the prevalence of B. pseudomallei in the environment, intensive sampling of soil and fresh water within Ocean Park, especially around marine mammal facilities, was done. This resulted in isolation rates of about 1.4% in soil, 14% in fresh water, and 0% in seawater. For the rest of Hong Kong, isolation rates have been about 1.5% in soil and 14% in fresh water.33 These environmental isolation rates are similar both within and outside Ocean Park. This suggests that the organism is distributed relatively evenly throughout Hong Kong and is not concentrated at this facility. The prevalence of B. pseudomallei in soil and water in Hong Kong is considered low when compared to northeastern Thailand, which has both a high soil isolation rate of 50% to 68% and a high prevalence of human melioidosis.44,56,62
Subjective observations at Ocean Park revealed that melioidosis cases in marine mammals occurred shortly after severe tropical cyclones or typhoons, and the stronger the typhoon, the greater the risk of developing disease. This observation was supported by the isolation of B. pseudomallei from moisture collected from strong winds during typhoons. Findings from environmental isolation studies at Ocean Park, although still circumstantial, suggest that strong winds and rain might carry and move B. pseudomallei from distant sites to expose susceptible animals. The organism has only been isolated (from wind and rainwater) within marine mammal facilities at Ocean Park during inclement weather, and it has not been isolated from soil or water near these facilities. The human literature also reports similar associations; a cyclone in Western Australia was followed by the first cases of acute melioidosis in a previously nonendemic area.29
Humans and Other Animals
Clinical manifestations of melioidosis may be variable, thereby earning its nickname of the “great imitator.” Descriptions in humans include an acute “disseminated” septicemia with a short course, often resulting in death despite treatment; a pulmonary form; and a chronic form presenting with abscessation or granuloma formation in almost any organ system.16 However, it is believed that the majority of infections in people must be mild or asymptomatic because up to 49% of some populations in endemic areas have developed antibodies to B. pseudomallei.32 Patients who develop septicemic melioidosis are acutely critically ill, have reduced consciousness, are pyrexic and diarrheic, and have multiorgan involvement, typically of the lungs, liver, and spleen.5 The pulmonary form may develop either as a primary pneumonitis secondary to septicemia or more chronically as a cavitating pneumonia. Localized infections of other organs and organ systems include skin, subcutis, muscles, joints, glandular tissue, lymphatic tissue, bone, genitalia, urinary tract, and central nervous system.
In other animals, acute septicemia, pulmonary lesions, and abscessation in virtually any organ system have also been reported. Abscesses and microabscesses, which may be described as “nodular,” are mainly distributed in the liver, spleen, lymph nodes, and lungs of affected animals. The lesions are often enclosed within a fibrous capsule.35 Lameness resulting from suppurative polyarthritis is relatively common,12,50,51 and nodular purulent lymphangitis causing lameness has also been described.7 Other presentations include mastitis, especially in goats and sheep12,38,39; orchitis and periorchitis7,21; and aortic aneurysms.12 Transplacental infection causing abortions has been described.30,52 Asymptomatic infection of livestock in Australia appears to be a common outcome, with melioidosis diagnosed by isolating B. pseudomallei from abscesses during routine meat inspection.31,51
Almost all cetaceans present with acute septicemic melioidosis. The clinical presentation of these cases was nonspecific and included an acute onset of anorexia, pyrexia, and lethargy. Pyrexia appears to be a consistent early indicator of disease. Dyspnea and extreme dullness with inactivity were often observed terminally. Occasionally the cetacean was found dead in the pool or presented at a terminal stage. After the onset of clinical signs, 29 animals died in about 5 days (mean). Hematology usually demonstrated an initial neutrophilia with a left shift, with or without a concurrent leukocytosis, and often leukopenia before death. Liver enzymes (alanine transaminase, aspartate transaminase, lactate dehydrogenase, gamma-glutamyltransferase) were often elevated and alkaline phosphatase was decreased on serum chemistries. The closest to the chronic localized form of melioidosis was in a bottlenose dolphin with osteomyelitis of the lumbar vertebrae and extensive abscessation around and erosion of the dorsal spinous processes of the tenth and eleventh thoracic vertebrae. This male dolphin had a 5-month history of a decreased appetite, weight loss, and later reduced mobility of the spine. He eventually succumbed to disseminated septicemic melioidosis.
In pinnipeds, as in cetaceans, most cases at Ocean Park presented as an acute septicemia. The clinical presentation was nonspecific and included reluctance or refusal to perform, lethargy and dullness, tachypnea, diarrhea, and partial to complete anorexia. Pyrexia was present in all eight cases for which body temperature was obtained and recorded. The mean time to death after the first clinical signs for most pinnipeds was about 5 days (range, 1-23 days). Hematologic and serum biochemical findings were similar to those in cetaceans. One atypical case was a female California sea lion (Zalophus californianus) that displayed signs of localized melioidosis. She presented 41 days before death with inappetence, dullness, and abscessation (thick, yellow/cream-colored purulent material cultured positive for B. pseudomallei) and cellulitis of the mammary glands and surrounding subcutis. Despite treatment, which included surgical drainage, debridement, placement of indwelling drains, flushing of the site, and fluid plus antimicrobial therapy, the sea lion developed septicemic melioidosis and died. Hematology and serum chemistry revealed a neutrophilia with a left shift and an increasing leukocytosis throughout the sea lion’s illness, with progressive anemia and hyperglobinemia. The duration of malaise and the development of extensive pockets of abscessation in the mammary tissue and subcutis suggest chronic localized melioidosis.