Locally Acquired Melioidosis Linked to Environment - Mississippi, 2020-2023.

Melioidosis, caused by Burkholderia pseudomallei, is a rare but potentially fatal bacterial disease endemic to tropical and subtropical regions worldwide. It is typically acquired through contact with contaminated soil or fresh water. Before this investigation, B. pseudomallei was not known to have been isolated from the environment in the continental United States. Here, we report on three patients living in the same Mississippi Gulf Coast county who presented with melioidosis within a 3-year period. They were infected by the same Western Hemisphere B. pseudomallei strain that was discovered in three environmental samples collected from the property of one of the patients. These findings indicate local acquisition of melioidosis from the environment in the Mississippi Gulf Coast region.

Multistate Outbreak of Melioidosis Associated with Imported Aromatherapy Spray.

Melioidosis, caused by the bacterium Burkholderia pseudomallei, is an uncommon infection that is typically associated with exposure to soil and water in tropical and subtropical environments. It is rarely diagnosed in the continental United States. Patients with melioidosis in the United States commonly report travel to regions where melioidosis is endemic. We report a cluster of four non-travel-associated cases of melioidosis in Georgia, Kansas, Minnesota, and Texas. These cases were caused by the same strain of B. pseudomallei that was linked to an aromatherapy spray product imported from a melioidosis-endemic area.

Plasma Metabolomics Reveals Distinct Biological and Diagnostic Signatures for Melioidosis.

Rationale: The global burden of sepsis is greatest in low-resource settings. Melioidosis, infection with the gram-negative bacterium Burkholderia pseudomallei, is a frequent cause of fatal sepsis in endemic tropical regions such as Southeast Asia. Objectives: To investigate whether plasma metabolomics would identify biological pathways specific to melioidosis and yield clinically meaningful biomarkers. Methods: Using a comprehensive approach, differential enrichment of plasma metabolites and pathways was systematically evaluated in individuals selected from a prospective cohort of patients hospitalized in rural Thailand with infection. Statistical and bioinformatics methods were used to distinguish metabolomic features and processes specific to patients with melioidosis and between fatal and nonfatal cases. Measurements and Main Results: Metabolomic profiling and pathway enrichment analysis of plasma samples from patients with melioidosis (n = 175) and nonmelioidosis infections (n = 75) revealed a distinct immuno-metabolic state among patients with melioidosis, as suggested by excessive tryptophan catabolism in the kynurenine pathway and significantly increased levels of sphingomyelins and ceramide species. We derived a 12-metabolite classifier to distinguish melioidosis from other infections, yielding an area under the receiver operating characteristic curve of 0.87 in a second validation set of patients. Melioidosis nonsurvivors (n = 94) had a significantly disturbed metabolome compared with survivors (n = 81), with increased leucine, isoleucine, and valine metabolism, and elevated circulating free fatty acids and acylcarnitines. A limited eight-metabolite panel showed promise as an early prognosticator of mortality in melioidosis. Conclusions: Melioidosis induces a distinct metabolomic state that can be examined to distinguish underlying pathophysiological mechanisms associated with death. A 12-metabolite signature accurately differentiates melioidosis from other infections and may have diagnostic applications.

Layering vaccination with antibiotic therapy results in protection and clearance of Burkholderia pseudomallei in Balb/c mice.

Melioidosis is a disease that is difficult to treat due to the causative organism, Burkholderia pseudomallei being inherently antibiotic resistant and it having the ability to invade, survive, and replicate in an intracellular environment. Combination therapy approaches are routinely being evaluated in animal models with the aim of improving the level of protection and clearance of colonizing bacteria detected. In this study, a subunit vaccine layered with the antibiotic finafloxacin was evaluated in vivo against an inhalational infection with B. pseudomallei in Balb/c mice. Groups of mice vaccinated, infected, and euthanized at antibiotic initiation had a reduced bacterial load compared to those that had not been immunized. In addition, the subunit vaccine provided a synergistic effect when it was delivered with a CpG ODN and finafloxacin was initiated at 48 h post-challenge. Vaccination was also shown to improve the outcome, in a composite measure of survival and clearance. In summary, layering a subunit vaccine with the antibiotic finafloxacin is a promising therapeutic alternative for use in the treatment of B. pseudomallei infections.

Flagellin-modulated inflammasome pathways characterize the human alveolar macrophage response to Burkholderia pseudomallei, a lung-tropic pathogen.

Melioidosis is an emerging tropical infection caused by inhalation, inoculation, or ingestion of the flagellated, facultatively intracellular pathogen Burkholderia pseudomallei. The melioidosis case fatality rate is often high, and pneumonia, the most common presentation, doubles the risk of death. The alveolar macrophage is a sentinel pulmonary host defense cell, but the human alveolar macrophage in B. pseudomallei infection has never been studied. The objective of this study was to investigate the host-pathogen interaction of B. pseudomallei infection with the human alveolar macrophage and to determine the role of flagellin in modulating inflammasome-mediated pathways. We found that B. pseudomallei infects primary human alveolar macrophages but is gradually restricted in the setting of concurrent cell death. Electron microscopy revealed cytosolic bacteria undergoing division, indicating that B. pseudomallei likely escapes the alveolar macrophage phagosome and may replicate in the cytosol, where it triggers immune responses. In paired human blood monocytes, uptake and intracellular restriction of B. pseudomallei are similar to those observed in alveolar macrophages, but cell death is reduced. The alveolar macrophage cytokine response to B. pseudomallei is characterized by marked interleukin (IL)-18 secretion compared to monocytes. Both cytotoxicity and IL-18 secretion in alveolar macrophages are partially flagellin dependent. However, the proportion of IL-18 release that is driven by flagellin is greater in alveolar macrophages than in monocytes. These findings suggest differential flagellin-mediated inflammasome pathway activation in the human alveolar macrophage response to B. pseudomallei infection and expand our understanding of intracellular pathogen recognition by this unique innate immune lung cell.

Extracorporeal Membrane Oxygenation-Dependent Fulminant Melioidosis From Caspase 4 Mutation Reversed by Interferon Gamma Therapy.

We describe bedside-to-bench immunological and genetic elucidation of defective pyroptosis attributable to novel caspase 4 defect mediating pathogen-triggered inflammatory programmed cell death, in the setting of severe pneumonia and abscess-forming melioidosis in an overtly healthy host failing to clear Burkholderia pseudomallei infection, and how targeted adjunctive biological therapy led to a successful outcome.

Burkholderia pseudomallei Bacteria in Ornamental Fish Tanks, Vientiane, Laos, 2023.

In 2019, a melioidosis case in Maryland, USA, was shown to have been acquired from an ornamental fish tank contaminated with Burkholderia pseudomallei bacteria, likely derived from Southeast Asia. We investigated the presence of B. pseudomallei in ornamental fish tanks in the endemic area of Vientiane, Laos.

Melioidosis in Patients with COVID-19 Exposed to Contaminated Tap Water, Thailand, 2021.

In September 2021, a total of 25 patients diagnosed with COVID-19 developed acute melioidosis after (median 7 days) admission to a COVID-19 field hospital in Thailand. Eight nonpotable tap water samples and 6 soil samples were culture-positive for Burkholderia pseudomallei. Genomic analysis suggested contaminated tap water as the likely cause of illness.

N-acetyl-ß-hexosaminidase activity is important for chitooligosaccharide metabolism and biofilm formation in Burkholderia pseudomallei.

Burkholderia pseudomallei is a saprophytic Gram-negative bacillus that can cause the disease melioidosis. Although B. pseudomallei is a recognised member of terrestrial soil microbiomes, little is known about its contribution to the saprophytic degradation of polysaccharides within its niche. For example, while chitin is predicted to be abundant within terrestrial soils the chitinolytic capacity of B. pseudomallei is yet to be defined. This study identifies and characterises a putative glycoside hydrolase, bpsl0500, which is expressed by B. pseudomallei K96243. Recombinant BPSL0500 was found to exhibit activity against substrate analogues and GlcNAc disaccharides relevant to chitinolytic N-acetyl-ß-d-hexosaminidases. In B. pseudomallei, bpsl0500 was found to be essential for both N-acetyl-ß-d-hexosaminidase activity and chitooligosaccharide metabolism. Furthermore, bpsl0500 was also observed to significantly affect biofilm deposition. These observations led to the identification of BPSL0500 activity against model disaccharide linkages that are present in biofilm exopolysaccharides, a feature that has not yet been described for chitinolytic enzymes. The results in this study indicate that chitinolytic N-acetyl-ß-d-hexosaminidases like bpsl0500 may facilitate biofilm disruption as well as chitin assimilation, providing dual functionality for saprophytic bacteria such as B. pseudomallei within the competitive soil microbiome.

Synthesis of BLF1-containing trimethyl chitosan nanoparticles and evaluation of its immunogenicity and protection in syrian mice by oral and subcutaneous injections.

The bacterium Burkholderia pseudomallei is the cause of melioidosis infectious disease. In this bacterium, the BLF1 protein wide inhibits the synthesis of proteins in human cells. This disease is reported to cause a death rate of 40% in some parts of the world. Currently, no effective vaccine is available against this bacterial infection. In this study, therefore, a Nano vaccine was synthesized based on the trimethyl chitosan (TMC) polymer containing the BLF1 recombinant protein, and its immunogenicity and protection in Syrian mice were evaluated by oral and subcutaneous injections. The BLF1 recombinant protein expression was induced in Escherichia coli Bl21 (DE3) and purified by the affinity chromatography technique. Recombinant protein-containing nanoparticles (NPs) were then synthesized by the ionotropic gelation method. After oral and subcutaneous injections, antibody titration was assessed by the indirect ELISA assay. Finally, murine groups were challenged using the BLF1 toxin. The results indicated that the immune system showed more antibody titration in subcutaneous injection than in the oral form. However, the results were reversed in the challenge results, and the survival rate was more significant in the oral injection.

Paediatric melioidosis.

Melioidosis is a tropical infectious disease caused by the saprophytic gram-negative bacterium Burkholderia pseudomallei. Despite the infection being endemic in southeast Asia and northern Australia, the broad clinical presentations and diagnostic difficulties limit its early detection, particularly in children. Melioidosis more commonly affects the immunocompromised and adults. Melioidosis is increasingly being diagnosed around the world and whole-genome sequencing indicates that these cases are not linked with travel to endemic areas. Research has concentrated on the adult population with limited experience reported in the care of this uncommon, but potentially fatal condition in children presenting with bacteraemia and pneumonia.

Development of a novel sequence based real-time PCR assay for specific and sensitive detection of Burkholderia pseudomallei in clinical and environmental matrices.

BACKGROUND: Melioidosis, caused by the category B biothreat agent Burkholderia pseudomallei, is a disease with a high mortality rate and requires an immediate culture-independent diagnosis for effective disease management. In this study, we developed a highly sensitive qPCR assay for specific detection of Burkholderia pseudomallei and melioidosis disease diagnosis based on a novel target sequence. METHODS: An extensive in-silico analysis was done to identify a novel and highly conserved sequence for developing a qPCR assay. The specificity of the developed assay was analyzed with 65 different bacterial cultures, and the analytical sensitivity of the assay was determined with the purified genomic DNA of B. pseudomallei. The applicability of the assay for B. pseudomallei detection in clinical and environmental matrices was evaluated by spiking B. pseudomallei cells in the blood, urine, soil, and water along with suitable internal controls. RESULTS: A novel 85-nucleotide-long sequence was identified using in-silico tools and employed for the development of the highly sensitive and specific quantitative real-time PCR assay S664. The assay S664 was found to be highly specific when evaluated with 65 different bacterial cultures related and non-related to B. pseudomallei. The assay was found to be highly sensitive, with a detection limit of 3 B. pseudomallei genome equivalent copies per qPCR reaction. The detection limit in clinical matrices was found to be 5 × 102 CFU/mL for both human blood and urine. In environmental matrices, the detection limit was found to be 5 × 101 CFU/mL of river water and 2 × 103 CFU/gm of paddy field soil. CONCLUSIONS: The findings of the present study suggest that the developed assay S664 along with suitable internal controls has a huge diagnostic potential and can be successfully employed for specific, sensitive, and rapid molecular detection of B. pseudomallei in various clinical and environmental matrices.

Isolation of Burkholderia pseudomallei from a goat in New Caledonia: implications for animal and human health monitoring and serological tool comparison.

BACKGROUND: Melioidosis is a serious bacterial infection caused by Burkholderia pseudomallei, a gram-negative bacterium commonly found in soil and water. It can affect both humans and animals, and is endemic in regions such as Southeast Asia and Northern Australia. In recent years, there have been reports of an emergence of human melioidosis in other areas, including New Caledonia. RESULTS: During standard laboratory analysis in New Caledonia in 2021, a strain of B. pseudomallei was isolated from a goat. The strain was characterized using both MLST and WGS techniques and was found to cluster with previously described local human strains from the area. In parallel, several serological tests (CFT, ELISA, Luminex (Hcp1, GroEL, BPSS1840), arrays assay and a latex agglutination test) were performed on animals from the farm where the goat originated, and/or from three other neighboring farms. Using two commercial ELISA kits, seropositive animals were found only on the farm where the infected goat originated and tests based on recombinant proteins confirmed the usefulness of the Hcp1 protein for the diagnosis of melioidosis in animals. CONCLUSIONS: Despite the regular reports of human cases, this is the first confirmed case of melioidosis in an animal in New Caledonia. These results confirm the presence of the bacterium in the region and highlight the importance of vigilance for both animal and human health. It is critical that all health partners, including breeders, veterinarians, and biologists, work together to monitor and prevent the spread of the disease.

Function of Burkholderia pseudomallei RpoS and RpoN2 in bacterial invasion, intracellular survival, and multinucleated giant cell formation in mouse macrophage cell line.

Melioidosis, a human infectious disease with a high mortality rate in many tropical countries, is caused by the pathogen Burkholderia pseudomallei (B. pseudomallei). The function of the B. pseudomallei sigma S (RpoS) transcription factor in survival during the stationary growth phase and conditions of oxidative stress is well documented. Besides the rpoS, bioinformatics analysis of B. pseudomallei genome showed the existence of two rpoN genes, named rpoN1 and rpoN2. In this study, by using the mouse macrophage cell line RAW264.7 as a model of infection, the involvement of B. pseudomallei RpoS and RpoN2 in the invasion, intracellular survival leading to the reduction in multinucleated giant cell (MNGC) formation of RAW264.7 cell line were illustrated. We have demonstrated that the MNGC formation of RAW264.7 cell was dependent on a certain number of intracellular bacteria (at least 5 × 104). In addition, the same MNGC formation (15%) observed in RAW264.7 cells infected with either B. pseudomallei wild type with multiplicity of infection (MOI) 2 or RpoN2 mutant (∆rpoN2) with MOI 10 or RpoS mutant (∆rpoS) with MOI 100. The role of B. pseudomallei RpoS and RpoN2 in the regulation of type III secretion system on bipB-bipC gene expression was also illustrated in this study.

Diabetes and Infectious Diseases with a Focus on Melioidosis.

Diabetes mellitus (DM) leads to impaired innate and adaptive immune responses. This renders individuals with DM highly susceptible to microbial infections such as COVID-19, tuberculosis and melioidosis. Melioidosis is a tropical disease caused by the bacterial pathogen Burkholderia pseudomallei, where diabetes is consistently reported as the most significant risk factor associated with the disease. Type-2 diabetes is observed in 39% of melioidosis patients where the risk of infection is 13-fold higher than non-diabetic individuals. B. pseudomallei is found in the environment and is an opportunistic pathogen in humans, often exhibiting severe clinical manifestations in immunocompromised patients. The pathophysiology of diabetes significantly affects the host immune responses that play a critical role in fighting the infection, such as leukocyte and neutrophil impairment, macrophage and monocyte inhibition and natural killer cell dysfunction. These defects result in delayed recruitment as well as activation of immune cells to target the invading B. pseudomallei. This provides an advantage for the pathogen to survive and adapt within the immunocompromised diabetic patients. Nevertheless, knowledge gaps on diabetes-infectious disease comorbidity, in particular, melioidosis-diabetes comorbidity, need to be filled to fully understand the dysfunctional host immune responses and adaptation of the pathogen under diabetic conditions to guide therapeutic options.

The Predictive Factors Associated with In-Hospital Mortality of Melioidosis: A Cohort Study.

Background and Objectives: Melioidosis is an infectious disease caused by Burkholderia pseudomallei, and it has a wide range of clinical symptoms. It is endemic in tropical areas, including Southeast Asia. Despite the availability of effective treatment, the mortality rate is still high, especially in patients presenting with septic shock. The aim of this study was to determine and explore clinical characteristics, microbiology, treatment outcomes, and factors associated with in-hospital mortality which could predict prognosis and provide a guide for future treatment. Materials and Methods: The population in this retrospective cohort study included all 262 patients with a diagnosis of melioidosis who were hospitalized at Surin Hospital, Surin, Thailand, from April 2014 to March 2017. We included patients older than 15 years with a positive culture for B. pseudomallei. Data regarding the clinical characteristics, microbiology, and treatment outcomes of the patients were collected and analyzed. The patients were divided into two groups dependent on outcome, specifically non-survival and survival. Logistic regression was performed to determine the risk factors associated with in-hospital mortality. Results: Out of the 262 patients with melioidosis during the study period, 117 (44.7%) patients died. The mean age was 57.2 ± 14.4 years, and 193 (73.7%) patients were male. The most common comorbidity was diabetes (123, 46.9%), followed by chronic kidney disease (35, 13.4%) and chronic liver disease (31, 11.8%). Four risk factors were found to be associated with in-hospital mortality, including age (adjusted odds ratio (aOR) 1.04, 95%CI: 1.01-1.07), respiration rate (aOR 1.18, 95%CI: 1.06-1.32), abnormal chest X-ray finding (aOR 4.79, 95%CI: 1.98-11.59), and bicarbonate levels (CO2) (aOR 0.92, 95%CI: 0.85-0.99). Conclusions: Our study identified age, respiration rate, abnormal chest X-ray finding, and CO2 levels are predictive factors associated with in-hospital mortality in melioidosis patients. Physicians should be aware of these factors, have access to aggressive treatment options, and closely monitor patients with these risk factors.

The Arabinose 5-Phosphate Isomerase KdsD Is Required for Virulence in Burkholderia pseudomallei.

Burkholderia pseudomallei is the causative agent of melioidosis, which is endemic primarily in Southeast Asia and northern Australia but is increasingly being seen in other tropical and subtropical regions of the world. Melioidosis is associated with high morbidity and mortality rates, which is mediated by the wide range of virulence factors encoded by B. pseudomallei. These virulence determinants include surface polysaccharides such as lipopolysaccharide (LPS) and capsular polysaccharides (CPS). Here, we investigated a predicted arabinose-5-phosphate isomerase (API) similar to KdsD in B. pseudomallei strain K96243. KdsD is required for the production of the highly conserved 3-deoxy-d-manno-octulosonic acid (Kdo), a key sugar in the core region of LPS. Recombinant KdsD was expressed and purified, and API activity was determined. Although a putative API paralogue (KpsF) is also predicted to be encoded, the deletion of kdsD resulted in growth defects, loss of motility, reduced survival in RAW 264.7 murine macrophages, and attenuation in a BALB/c mouse model of melioidosis. Suppressor mutations were observed during a phenotypic screen for motility, revealing single nucleotide polymorphisms or indels located in the poorly understood CPS type IV cluster. Crucially, suppressor mutations did not result in reversion of attenuation in vivo. This study demonstrates the importance of KdsD for B. pseudomallei virulence and highlights further the complex nature of the polysaccharides it produces. IMPORTANCE The intrinsic resistance of B. pseudomallei to many antibiotics complicates treatment. This opportunistic pathogen possesses a wide range of virulence factors, resulting in severe and potentially fatal disease. Virulence factors as targets for drug development offer an alternative approach to combat pathogenic bacteria. Prior to initiating early drug discovery approaches, it is important to demonstrate that disruption of the target gene will prevent the development of disease. This study highlights the fact that KdsD is crucial for virulence of B. pseudomallei in an animal model of infection and provides supportive phenotypic characterization that builds a foundation for future therapeutic development.

Presence of Burkholderia pseudomallei in Soil, Nigeria, 2019.

Melioidosis, caused by the soil-dwelling bacterium Burkholderia pseudomallei, is predicted to be endemic in Nigeria but is only occasionally reported. This report documents the systematic identification of the presence of B. pseudomallei and B. thailandensis in the soil across multiple states in Nigeria.

Burkholderia pseudomallei Laboratory Exposure, Arizona, USA.

We describe an incidental Burkholderia pseudomallei laboratory exposure in Arizona, USA. Because melioidosis cases are increasing in the United States and B. pseudomallei reservoirs have been discovered in the Gulf Coast Region, US laboratory staff could be at increased risk for B. pseudomallei exposure.

Burkholderia thailandensis Isolated from the Environment, United States.

Burkholderia thailandensis, an opportunistic pathogen found in the environment, is a bacterium closely related to B. pseudomallei, the cause of melioidosis. Human B. thailandensis infections are uncommon. We isolated B. thailandensis from water in Texas and Puerto Rico and soil in Mississippi in the United States, demonstrating a potential public health risk.