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PubMed1989

Identification of Pseudomonas pseudomallei in clinical practice: use of simple screening tests and API 20NE.

The API 20NE kit and a simple screening system involving Gram's stain, the oxidase reaction, colistin and gentamicin resistance, and colonial characteristics on a differential agar medium, were used to test 400 strains of Pseudomonas pseudomallei. The API kit identified 390 (97.5%) strains correctly on first testing and all but one of the remainder on second testing. Only one strain was initially misidentified (as Ps cepacia). The screening system was 100% accurate in identifying Ps pseudomallei. In non-endemic areas the API 20NE kit may be used to identify sporadic imported strains of Ps pseudomallei. Such kits may also help to delineate the geographical distribution of melioidosis. In endemic areas the screening tests described offer a cheap, simple, and accurate means of presumptively identifying Ps pseudomallei from clinical specimens.

Bacteriological Techniques↗

PubMed2002

Strategies for PCR based detection of Burkholderia pseudomallei DNA in paraffin wax embedded tissues.

Recently, several cases of melioidosis imported to Europe have been reported. The diagnosis of the acute or chronic infection remains challenging. This report describes an optimised protocol for fast and reliable DNA preparation for use in two different polymerase chain reaction (PCR) assays, namely: (1) a seminested PCR assay targeting a genus specific sequence of the ribosomal protein subunit 21 (rpsU) gene and (2) a nested PCR assay targeting the gene encoding the filament forming flagellin (fliC). Various strains of Burkholderia spp, strains of closely related genera, and spleen tissue samples of experimentally infected mice were investigated. The combination of PCR and sequencing of the amplicons resulted in high sensitivity and specificity. These procedures may allow rapid, sensitive, and reliable detection of B pseudomallei DNA in routinely formalin fixed and paraffin wax embedded samples, thus providing a safe diagnostic tool and avoiding the cultivation of a risk group 3 agent. In addition, this method could be useful for retrospective histopathological investigations.

Animals↗

PubMed2006

In vitro antimicrobial activity of natural toxins and animal venoms tested against Burkholderia pseudomallei.

BACKGROUND: Burkholderia pseudomallei are the causative agent of melioidosis. Increasing resistance of the disease to antibiotics is a severe problem in treatment regime and has led to intensification of the search for new drugs. Antimicrobial peptides are the most ubiquitous in nature as part of the innate immune system and host defense mechanism. METHODS: Here, we investigated a group of venoms (snakes, scorpions and honey bee venoms) for antimicrobial properties against two strains of Gram-negative bacteria Burkholderia pseudomallei by using disc-diffusion assay for in vitro susceptibility testing. The antibacterial activities of the venoms were compared with that of the isolated L-amino acid oxidase (LAAO) and phospholipase A2 (PLA2s) enzymes. MICs were determined using broth dilution method. Bacterial growth was assessed by measurement of optical density at the lowest dilutions (MIC 0.25 mg/ml). The cell viability was measured using tetrazolium salts (XTT) based cytotoxic assay. RESULTS: The studied venoms showed high antimicrobial activity. The venoms of C. adamanteus, Daboia russelli russelli, A. halys, P. australis, B. candidus and P. guttata were equally as effective as Chloramphenicol and Ceftazidime (30 microg/disc). Among those tested, phospholipase A2 enzymes (crotoxin B and daboiatoxin) showed the most potent antibacterial activity against Gram-negative (TES) bacteria. Naturally occurring venom peptides and phospholipase A2 proved to possess highly potent antimicrobial activity against Burkholderia pseudomallei. The XTT-assay results showed that the cell survival decreased with increasing concentrations (0.05-10 mg/mL) of Crotalus adamanteus venom, with no effect on the cell viability evident at 0.5 mg/mL. CONCLUSION: This antibacterial profile of snake venoms reported herein will be useful in the search for potential antibacterial agents against drug resistant microorganisms like B. pseudomallei.

Animals↗

PubMed2004

Burkholderia pseudomallei: a case report of a human infection in Ceará, Brazil.

Burkholderia pseudomallei has rarely been isolated from environmental and clinical specimens in South America, particularly, in Brazil. This report describes a case of melioidosis with fulminant sepsis in a 10 year old boy, from rural area, in Tejuçuoca, State of Ceará, Brazil. Blood samples were positive and, through the analysis of results from biochemical tests and of drugs susceptibility profile, identified this gram-negative bacillus as B. pseudomallei. The contamination source remains obscure in this case, as soil and water tanks samples submitted to microbiological analyses did not indicate the presence of B. pseudomallei.

Brazil↗

PubMedDate not supplied

PCR-based identification of Burkholderia pseudomallei.

DNA amplification techniques are being used increasingly in clinical laboratories to confirm the identity of medically important bacteria. A PCR-based identification method has been in use in our centre for 10 years for Burkholderia pseudomallei and was used to confirm the identity of bacteria isolated from cases of melioidosis in Ceará since 2003. This particular method has been used as a reference standard for less discriminatory methods. In this study we evaluated three PCR-based methods of B. pseudomallei identification and used DNA sequencing to resolve discrepancies between PCR-based results and phenotypic identification methods. The established semi-nested PCR protocol for B. pseudomallei 16-23s spacer region produced a consistent negative result for one of our 100 test isolates (BCC #99), but correctly identified all 71 other B. pseudomallei isolates tested. Anomalous sequence variation was detected at the inner, reverse primer binding site for this method. PCR methods were developed for detection of two other B. pseudomallei bacterial metabolic genes. The conventional lpxO PCR protocol had a sensitivity of 0.89 and a specificity of 1.00, while a real-time lpxO protocol performed even better with sensitivity and specificity of 1.00, and 1.00. This method identified all B. pseudomallei isolates including the PCR-negative discrepant isolate. The phaC PCR protocol detected the gene in all B. pseudomallei and all but three B. cepacia isolates, making this method unsuitable for PCR-based identification of B. pseudomallei. This experience with PCR-based B. pseudomallei identification methods indicates that single PCR targets should be used with caution for identification of these bacteria, and need to be interpreted alongside phenotypic and alternative molecular methods such as gene sequencing.

Bacterial Typing Techniques↗

PubMed2006

Phenotypic characterization of three clinical isolates of Burkholderia pseudomallei in Ceará, Brazil.

Burkholderia pseudomallei, the causative agent of melioidosis was found in a small cluster of cases in Tejuçuoca, Ceará, Brazil. Tests were carried out to determine its phenotypic characteristics: colony morphology on Ashdown agar and MacConkey agar, biochemical profile in conventional biochemical tests and API 20NE, arabinose assimilation and susceptibility testing by disk diffusion, comparing with data in the literature. This study confirms the presence of B. pseudomallei in Brazil and describes its characteristics.

Adolescent↗

PubMed2003

Melioidotic septic arthritis and its risk factors.

BACKGROUND: Melioidotic septic arthritis is an infection caused by the gram-negative bacillus Burkholderia pseudomallei. It is commonly found in Northeast Thailand. The goal of our study was to identify specific characteristics of patients with melioidotic septic arthritis by comparing them with patients with non-melioidotic septic arthritis and to describe the results of treatment of melioidotic septic arthritis. METHODS: We conducted a retrospective study of seventy-seven patients with septic arthritis who were treated in our hospital over a period of four years. Twenty-five of the patients had melioidotic septic arthritis, and fifty-two had non-melioidotic septic arthritis. Univariate and multivariate analyses were conducted to identify the risk factors for melioidotic septic arthritis, and the clinical course of the twenty-five patients with melioidotic septic arthritis was followed until the infection resolved. RESULTS: Patients with melioidotic septic arthritis differed significantly (p = 0.002 ) from those with non-melioidotic septic arthritis with regard to the frequency of diabetes mellitus and of involvement of an upper-extremity joint. The odds ratio that melioidosis was the cause of the infection was 15.7 (95% confidence interval, 4.5 to 55.6) in a patient with diabetes mellitus and 4.51 (95% confidence interval, 1.04 to 19.65) in a patient with involvement of an upper-extremity joint. Twenty-two of the twenty-five patients with melioidotic septic arthritis responded to treatment, which consisted of six months of antibiotic therapy combined with needle aspiration, as well as surgical drainage of the affected joint when necessary (sixteen patients). CONCLUSIONS: A diagnosis of melioidotic septic arthritis should be considered when septic arthritis is seen in an individual who is indigenous to or has recently visited Southeast Asia. The infection is more likely to be melioidotic septic arthritis if it involves an upper-extremity joint and if the patient has diabetes mellitus.

Adult↗

PubMed1981

Demonstration of human antibodies to Pseudomonas pseudomallei by indirect fluorescent antibody staining.

A rapid, reliable immunofluorescent assay for human antibodies to Pseudomonas pseudomallei was devised. The method was found to be sensitive and specific by inhibition tests and by tests for cross reactivity with antisera and cells of closely related pseudomonads other than Pseudomonas mallei. Preliminary studies suggest that the technique may be useful as a screening test for the detection of melioidosis and that demonstration of specific IgM by immunofluorescence may be of value in differentiating active from inactive infection.

Antibodies, Bacterial↗

PubMed1994

Sherlock Holmes and tropical medicine: a centennial appraisal.

Sir Arthur Conan Doyle incorporated an unidentified tropical disease as a murder weapon in the Sherlock Holmes story, "The Dying Detective," written in 1913. Documentary and circumstantial evidence suggests that the disease mentioned was melioidosis. The description of the newly identified disease occurred shortly before Doyle's death. Doyle's other works at the time reflect a consistent interest in tropical disease.

England↗

PubMed1996

Short report: Electron microscopic demonstration of extracellular structure of Burkholderia pseudomallei.

The occurrence of latency and relapse in human melioidosis suggests adaptations by Burkholderia pseudomallei that help to avoid the human immune response. Ruthenium red-stained preparations of bacterial cultures viewed by electron microscopy revealed three morphologically distinct variants; one with a very marked and another with a less electron-dense layer surrounding the cell wall, and a third variety devoid of such a structure. This structure may be attributable to a layer of polysaccharide, suggesting the presence of a glycocalyx that may aid in the survival of the organism during latency.

Burkholderia pseudomallei↗

PubMed1997

Speed of detection of Burkholderia pseudomallei in blood cultures and its correlation with the clinical outcome.

Burkholderia pseudomallei is a major cause of fatalities from nonhospital-acquired, gram-negative bacterial septicemia in northeastern part of Thailand. Rapid isolation of the bacterium is critical for diagnosis and treatment. Bacterial culture is currently the gold standard method for laboratory diagnosis of melioidosis. The present study describes the time to detection of B. pseudomallei in blood cultures using a BacT/Alert automated blood culture system, and the correlation between the speed of detection and the clinical outcome of the patients. Of 813 consecutive positive blood cultures, 75 blood cultures from 71 patients were positive for B. pseudomallei. The mean +/- SD time to detection of growth of B. pseudomallei was 23.9 +/- 14.9 hr (95% confidence interval = 20.4-27.5 hr). A total of 62.5% of the B. pseudomallei-positive cultures was detected within 24 hr of incubation, and 93.1% within 48 hr. Interestingly, fatalities occurred in 73.7% of those in which the bacterial growth was detected within the first 24 hr, as compared with only 40.9% in those with a time to detection of culture more than 24 hr (P = 0.012). The shorter time of detection of the bacterial growth in blood cultures may reflect a higher bacterial level in the patient at the time blood was taken, and may be responsible for the poor clinical outcome.

Adolescent↗

PubMed1999

Epidemiology of Burkholderia pseudomallei in Thailand.

The distribution of Burkholderia pseudomallei in soil collected from four regions of Thailand and the frequency of B. pseudomallei infections in patients attending government hospitals throughout Thailand in 1997 were surveyed. A total of 3,585 soil samples collected from 896 sites in four regions of Thailand were cultured for B. pseudomallei using selective enrichment broth and modified Ashdown's agar. The organism was recovered in 4.4%, 6.1%, 20.4%, and 5.9% of the soil samples collected from the northern, central, northeastern, and southern regions, respectively, of Thailand (P < 0.0001). Burkholderia pseudomallei was cultured from 50.1% of the sites in the northeastern region compared with 13.8%, 24.5%, and 18.4% in the northern, central, and southern regions, respectively (P < 0.0001). The infection rate in patients attending government hospitals in the northeastern region (137.9 per 100,000 inpatients) was significantly higher than those in the northern (18 per 100,000 inpatients), central (13.4 per 100,000 inpatients), and southern (14.4 per 100,000 inpatients) regions, respectively (P < 0.0001). It is suggested that melioidosis, which is endemic in Thailand, is associated with the presence of B. pseudomallei in soil.

Burkholderia pseudomallei↗

PubMed1999

Mediation of attachment of Burkholderia pseudomallei to human pharyngeal epithelial cells by the asialoganglioside GM1-GM2 receptor complex.

Melioidosis is the term given to any infection caused by Burkholderia pseudomallei. This bacteria is one of the important causative agents of life-threatening pulmonary infections in the tropical and subtropical areas. The initiation of respiratory infections is attachment of this bacteria to pharyngeal cells. The precise mechanism of attachment of B. pseudomallei is not known. In this study, we found that asialoganglioside GM1 at concentrations of 25, 12.5, and 5 microg/ml significantly decreased the attachment of B. pseudomallei strain Sp-186 in a dose-dependent manner. On the other hand, asialoganglioside GM2 decreased the attachment of B. pseudomallei, but only at a concentration of 25 microg/ml. At a concentration of 1 mg/ml, glucose, N-acetyl-galactosamine, and galactose caused a significant decrease in attachment. However, at concentrations of 250 microg/ml, no decrease in attachment was observed in B. pseudomallei treated with these carbohydrates. Mannose and fucose at concentrations of 1 mg/ml had no effects on the inhibition of attachment of B. pseudomallei. Four other isolates of B. pseudomallei showed a significant decrease in attachment after treatment with asialoganglioside GM1. We conclude that asialogangliosides GM1 and GM2 are part of the receptor complex for B. pseudomallei on human pharyngeal epithelial cells.

Bacterial Adhesion↗

PubMed1993

Medicine in tropical Australia.

In the unique environment of Australia's tropical north there are endemic diseases inherited from Gondwana, others introduced from the north and from Europe, and a wide range of particularly venomous animals. There is continuing disparity in morbidity and mortality between Aboriginal people and other Australians in tropical areas and elsewhere. This is being addressed by the National Aboriginal Health Strategy, which emphasises social, environmental and economic issues, as well as control and coordination of services by Aboriginal and Torres Strait Islander communities. While the re-introduction of malaria remains a potential threat, together with other infections, current diseases in tropical Australia are being better elucidated; melioidosis is now recognised as the commonest cause of fatal [corrected] community-acquired pneumonia in the Top End of the Northern Territory, and a new focus of scrub typhus has been found. Sexually transmitted diseases are an urgent issue, especially for Aboriginal communities, given the potential impact of the human immunodeficiency virus.

Animals↗

PubMed1998

Chromobacterium violaceum in tropical northern Australia.

Chromobacterium violaceum is an unusual cause of infection which may have fatality rates of over 60% when systemic illness occurs. We describe four patients infected with this organism who were successfully treated at Royal Darwin Hospital between 1991 and 1996. In tropical Australia, C. violaceum infection should be considered along with melioidosis in patients who present with a septic illness with skin lesions and/or organ abscesses after exposure to soil in the wet season.

Anti-Bacterial Agents↗

PubMed1975

Human-to-human transmission of Pseudomonas pseudomallei.

Melioidosis, the clinical manifestation of infection with Pseudomonas pseudomallei, has occurred infrequently in American citizens; almost all reported cases have been in Vietnam veterans, usually associated with respiratory disease. A Vietnam veteran from Mississippi developed chronic prostatitis, with no other clinical manifestations, during service in Vietnam, and P. pseudomallei was isolated from prostatic secretions 2 years after his return to the United States. The patient had had sexual contact with four women including his wife since his return from Vietnam. Vaginal and cervical cultures and serum samples were obtained from the four women, and serum samples and cultures of semen were obtained from the patient. Vaginal swabs and semen cultures were negative for P. pseudomallei. The patient and his wife had hemagglutination titers (greater than 640) diagnostic of P. pseudominallei infection. This occurrence of venereal transmission is the first report of person-to-person spread of P. pseudomallei infection.

Adult↗

PubMedDate not supplied

Fatty acid profile and acid phosphatase activity of fresh isolates of Pseudomonas pseudomallei.

Eighty-one fresh isolates of Pseudomonas pseudomallei from melioidosis patients were subjected to the analysis for the fatty acid composition by gas-liquid chromatography (GLC) and pH-dependent pattern of nonspecific phosphatase activity. All the test strains were identical in the GLC profile showing the three peaks of characteristic hydroxy acids (3-OH 14:0, 2-OH 16:0, 3-OH 16:0) and the two prominent peaks of cyclopropane acids (17:0 delta, 19:0 delta). They had also basically the same pH-dependent curves of the enzymatic activity with paranitrophenyl phosphate as substrate, showing two to three peaks or shoulders only in the acidic side of the curve. These two biochemical characteristics could differentiate P. pseudomallei distinctly from P. aeruginosa, but not from P. cepacia.

Acid Phosphatase↗

PubMedDate not supplied

Serosurveillance for Pseudomonas pseudomallei infection in Thailand.

A nation-wide survey was conducted to see the prevalence of serosensitivity to Pseudomonas pseudomallei antigens by indirect hemagglutination (IHA) and indirect immunofluorescent assay (IFA) for IgG and IgM. Serum samples were collected from blood donors in eight selected areas and bacteriologically confirmed melioidosis patients in Ubon Ratchathani province. The distribution patterns of antibody titers were compared among the survey areas with cut-off points set at 1:160 for IHA, 1:4 for IFA-IgM and 1:32 for IFA-IgG. These cut-off points were decided by ROC (Receiver Operating Characteristics) analysis. The specificity (% true negative reactions) of each serological test in the general population differed significantly among survey areas, possibly reflecting the extent of inapparent infection in each community. IFA was more successful than IHA in differentiating between negative from positive reactions. The survey classified the areas into endemic (Khon Kaen, Ubon Ratchathani), transported (Bangkok), and non-endemic (other provinces) types.

Antibodies, Bacterial↗