BACKGROUND

Melioidosis is an infectious disease caused by Burkholderia pseudomallei. It is endemic in the Philippines and is underreported. Of the reported cases, the most common comorbidity is diabetes mellitus. The increasing cases of antibiotic resistance and the relatively high mortality rate highlights the need for increased awareness among clinicians regarding this disease. We aim to report a case of Burkholderia pseudomallei resistant to trimethoprim/sulfamethoxazole (TMP-SMX), used in its eradication following initial intravenous therapy.

A 51-year-old male Filipino with poor health-seeking behavior came with generalized body weakness, weight loss, dysarthria, fever, cough, difficulty breathing, bloatedness, dysuria, joint pains, and bilateral lower extremity hyperpigmented macules for four months. He has diabetes mellitus and hypertension and is a mechanic by trade. Initial workups revealed hemoglobin A1c (HbA1c) of 14.7%, and urinalysis with bacteriuria. Imaging revealed bilateral pneumonia on chest xray, hepatosplenomegaly on whole abdomen ultrasound, and old cerebral infarcts on cranial computed tomography scan (CT scan). Empiric antibiotics for the impression of sepsis from community-acquired pneumonia and urinary tract infection were ertapenem and azithromycin. Upon isolation of Burkholderia pseudomallei from blood cultures, the team shifted to TMP-SMX and ceftazidime for initial therapy of melioidosis. Sensitivity showed resistance to TMP-SMX; hence the team revised the antimicrobials to four weeks of levofloxacin and ceftazidime. After eleven hospital days, the team sent the patient home, clinically improved. The team continued levofloxacin for eradication therapy for three months and the patient responded well.

Fever with multi-system involvement in a Filipino patient with diabetes mellitus with significant environmental risk factors, poor glycemic control, splenomegaly, and treatment failure with appropriate empiric antibiotic therapy should raise suspicion for melioidosis. It is paramount that antimicrobial resistance be detected and documented upon isolation of Burkholderia pseudomallei, given the high relapse rates and the need for a prolonged duration of treatment.

Melioidosis is an infection caused by Burkholderia pseudomallei known to be endemic in large portions of Asia, Sub-Sahara, and North Australia. Despite its endemicity in Malaysia, prompt diagnosis and subsequent treatment remain elusive especially in the more peripheral medical centres. This coupled with increasing risk to the population because of worsening climate crises renders early recognition and treatment more justifiable than ever. Here we present a case of melioidosis septic arthritis with systemic dissemination and discuss the factors involved in disease contraction, worsening prevalence, and diagnostic methods.
Melioidosis ; Arthritis, Infectious ; mviN protein, Burkholderia pseudomallei [Supplementary Concept]

Background: Melioidosis is a potentially fatal disease caused by Burkholderia pseudomallei. Over a century after its discovery, there seems to be a paucity of reported cases in the Philippines relative to other countries where it is found to be endemic. This suggests that the true burden of melioidosis in the country is not well-defined. The rarity of the disease, its protean clinical manifestations, and the lack of pathognomonic features pose a great diagnostic challenge. Furthermore, the proper recognition of the organism is an extreme necessity as it is intrinsically resistant to numerous antibiotics and requires specific long-term treatment. Case: This is a case of a 49-year-old Filipino diagnosed with a metastatic spinal disease from a primary thyroid carcinoma and underwent posterior spinal decompression and stabilization. Revision of instrumentation was done following identification of an implant loosening. During the interim, wound dehiscence and infection developed. The patient was readmitted and underwent debridement of the lumbosacral spine. Wound cultures all yielded growth of Burkholderia pseudomallei. The patient received meropenem and then trimethoprim-sulfamethoxazole with ciprofloxacin during the intensive and eradication phase, respectively. Erythrocyte sedimentation rate and C-reactive protein were monitored and a significant reduction in both values reflected a good therapeutic response. Conclusion This is a rare case of a deep surgical site infection caused by Burkholderia pseudomallei. It is known that melioidosis is a potentially fatal infection but is under-reported in the Philippines. At present, further epidemiological studies along with an increased level of awareness of melioidosis are greatly needed to help define the true burden of illness and optimize patient management following prompt recognition.
Melioidosis ; Philippines ; Burkholderia pseudomallei

Aims: Every year, an estimated 25 million tons of waste oil are produced worldwide, and the generation of waste oil is one of the biggest global environmental problems. The incorporation of oil as a substrate for lipase production has been studied and shown to have a positive impact on its production. Burkholderia sp. is one of the major lipase-producing bacteria with their ability in bioremediation of oil-contaminated soil. This study aims to compare the production of lipase by Burkholderia cenocepacia ST8 using waste cooking oil and unused cooking oil as feedstock. Methodology and results: The effect of different types of waste cooking oil (sunflower oil and palm oil) and concentration (1-3%) of waste cooking oil, agitation speed (100-400 rpm) and initial dissolved oxygen concentration (10-50%) on lipase production by B. cenocepacia ST8 under batch fermentation mode were investigated. The major fatty acids of which had been consumed were determined using gas chromatography. Results showed that 2% (v/v) of single used sunflower cooking oil produced the highest lipase activity of 138.86 U/mL with a productivity of 2.10 U/mL/h; agitation speed of 300 rpm produced the highest lipase activity of 183.56 U/mL with a productivity of 3.06 U/mL/h while 30% initial concentration of dissolved oxygen produced a lipase activity of 176.45 U/mL with a productivity of 2.94 U/mL/h. Oleic acid and linoleic acid were found to be the most consumed by B. cenocepacia ST8 among other fatty acids. Conclusion, significance and impact of study This study shows that 2% (v/v) single used sunflower cooking oil was the better type and optimum concentration of carbon source for the production of lipase by the fermentation of B. cenocepacia under 300 rpm and 30% initial concentration dissolved oxygen. The incorporation of 2% (v/v) single used sunflower cooking oil may be a great alternative to reduce the cost for the production of lipase as well as reducing the amount of waste oil generation.
Lipase ; Burkholderia cenocepacia ; Waste Management ; Biodegradation, Environmental

Aims: Burkholderia pseudomallei, the human pathogen that causes melioidosis, is intrinsically resistant towards a wide range of antibiotics and there have been reports of acquired resistance towards antibiotics used for melioidosis treatments. Antimicrobial peptides (AMP) such as bacteriocins are gaining the interests of researchers as alternative for treating infections caused by multidrug resistant bacteria. In this study, we aimed to identify Burkholderia spp. isolated from soil in Sarawak that possess the potential in inhibiting the growth of B. pseudomallei and to further characterize the antagonistic compound produced. Methodology and results: A total of 50 Burkholderia spp. isolates of environmental origin and two isolates of Ralstonia solanacearum were screened against five clinical isolates of B. pseudomallei using spot-on-lawn assay and flip streak method. Burkholderia stagnalis isolate K23/3 showed clear zones of inhibition (ZOI) in both preliminary tests. Cell-free supernatant (CFS) was obtained from B. stagnalis K23/3 broth culture and was tested via agar well diffusion assay (AWDA). The antagonistic compound secreted at the early log phase of the bacterial growth was shown to be stable in a wide range of temperatures and pH. Treatment with different enzymes revealed that it was sensitive towards proteinase K, suggesting that it is proteinaceous. The bacteriocin-like-substance (BLIS) was subjected to ammonium sulfate precipitation and sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). The SDS-PAGE gel was overlaid with indicator B. pseudomallei isolates where the active protein was shown to be less than 7.1 kDa. Conclusion, significance and impact of study Burkholderia stagnalis isolate K23/3 was able to secrete bacteriocin-like-substance (BLIS) that has the potential in biocontrol of B. pseudomallei in the environment or as potential treatment for melioidosis.
Bacteriocins ; Burkholderia ; Burkholderia pseudomallei

In recent years, selenium nanoparticles (SeNPs) have been widely used in many fields such as nanotechnology, biomedicine and environmental remediation due to their good electrical conductivity, photothermal properties and anticancer properties. In this study, the cell-free supernatant, whole cell and the cell-free extracts of the strain Cupriavidus sp. SHE were used to synthesize SeNPs, and several methods were applied to analyze the crystal structure and surface functional groups of the nanoparticles. Finally, Pseudomonas sp. PI1 (G⁺) and Escherichia coli BL21 (G⁻) were selected to investigate the antibacterial properties of SeNPs. Cell-free supernatant, whole cell and cell-free extracts of the strain could synthesize SeNPs. As for the cell-free supernatant, selenite concentration of 5 mmol/L and pH=7 were favorable for the synthesis of SeNPs. TEM images show that the average size of nanospheres synthesized by the supernatant was 196 nm. XRD analysis indicates the hexagonal crystals structure of SeNPs. FTIR and SDS-PAGE confirmed the proteins bound to the surfaces of SeNPs. SeNPs synthesized by cell-free supernatant showed no antimicrobial activities against Pseudomonas sp. PI1 and Escherichia coli BL21 (DE3). These results suggest that proteins played an important role in biotransformation of SeNPs in an eco-friendly process, and SeNPs synthesized in this study were non-toxic and biologically compatible, which might be applied in other fields in the future.
Anti-Bacterial Agents ; pharmacology ; Bacteria ; drug effects ; Cupriavidus ; metabolism ; Nanoparticles ; Selenious Acid ; analysis ; Selenium ; chemistry ; pharmacology

Aims: Biocontrol of fungal plant pathogens using beneficial microorganisms is a safer alternative over synthetic fungicides. PHP12 is a bacterial strain isolated from healthy oil palm rhizosphere and is closely related to the recently described Burkholderia stagnalis, a member of the Burkholderia cepacia complex. This study aimed to characterize the antifungal activity spectrum of PHP12 and identify the antifungal compounds produced by the strain. Methodology and results: The antifungal activity of PHP12 was characterized by growing fungal strains in the presence and absence of PHP12 and measuring the radius of the antifungal zone. PHP12 inhibited the growth of fungal pathogens including Ganoderma boninense, Curvularia oryzae, Phellinus noxius and Colletotrichum capsici. However, PHP12 did not inhibit the growth of Trichoderma asperellum, a known fungal biocontrol agent. The antifungal compounds of PHP12 were precipitated using ammonium sulfate and further purified with HPLC followed by identification using Liquid Chromatography Electrospray Ionization Tandem Mass Spectrometric (LC/ESI-MS). The LC/ESI-MS analysis showed the presence of an oligopeptide with a molecular weight of 1210.63 Da. The peptide consists of heavily modified amino acids that are linked by a hexose residue. Conclusion, significance and impact of study Although characteristics of the antifungal compounds are similar to other antifungal peptides from Burkholderia such as occidiofungin, there have been no reports of antifungal peptides from B. stagnalis with the corresponding molecular weight or fragmentation profile. The novelty of the compound, as well as its antifungal spectrum, makes PHP12 an interesting strain to be investigated further as a biocontrol agent.
Fungicides, Industrial ; Burkholderia cepacia complex--pathogenicity

Ralstonia solanacearum strain PRS-84 used in this study was isolated from diseased Pogostemon cablin plants in our previous study.The competent cells of R.solanacearum strain PRS-84 were transformed by electroporation with Tn5 transposon and then were plated on TTC agar plates containing kanamycin to select for kanamycin-resistant colonies.The detection of kanamycin-resistant gene in kanamycin-resistant colonies was performed by PCR.Further,the flanking fragments of Tn5 transposon insertion site in the mutants were amplified by inverse PCR,and the flanking fragments were sequenced and analyzed.The results indicated that the kanamycin-resistant colonies were obtained in the transformation experiment of R.solanacearum strain PRS-84 by electroporation with Tn5 transposon.A specific band of approximately 700 bp was amplified by PCR from kanamycin-resistant colonies.The flanking sequences of Tn5 transposon insertion site in the transformants were obtained by inverse PCR.After sequencing and sequence analysis of Tn5 transposon insertion site in mutants,we preliminarily speculated that the Tn5 transposon inserted in the typ A gene,rec O gene and gid A gene in three mutants,respectively.A random mutagenesis system of R.solanacearum strain PRS-84 by electroporation with Tn5 transposon has been established,and the Tn5 insertion mutants have been obtained.This study might facilitate the creation of mutant library and the discovery of the virulence gene of R.solanacearum isolated from P.cablin.
DNA Transposable Elements ; Electroporation ; Genes, Bacterial ; Mutagenesis, Insertional ; Pogostemon ; microbiology ; Ralstonia solanacearum ; genetics ; Virulence

OBJECTIVE: To evaluate the synergy of the Burkholderia signaling molecule cis-2-dodecenoic acid (BDSF) and fluconazole (FLU) or itraconazole (ITRA) against two azole-resistant C. albicans clinical isolates in vitro and in vivo. METHODS: Minimum inhibitory concentrations (MICs) of antibiotics against two azole-resistant C. albicans were measured by the checkerboard technique, E-test, and time-kill assay. In vivo antifungal synergy testing was performed on mice. Analysis of the relative gene expression levels of the strains was conducted by quantitative reverse-transcription polymerase chain reaction (qRT-PCR). RESULTS: BDSF showed highly synergistic effects in combination with FLU or ITRA with a fractional inhibitory concentration index of ⪕ 0.08. BDSF was not cytotoxic to normal human foreskin fibroblast cells at concentrations of up to 300 μg/mL. The qRT-PCR results showed that the combination of BDSF and FLU/ITRA significantly inhibits the expression of the efflux pump genes CDR1 and MDR1 via suppression of the transcription factors TAC1 and MRR1, respectively, when compared with FLU or ITRA alone. No dramatic difference in the mRNA expression levels of ERG1, ERG11, and UPC2 was found, which indicates that the drug combinations do not significantly interfere with UPC2-mediated ergosterol levels. In vivo experiments revealed that combination therapy can be an effective therapeutic approach to treat candidiasis. CONCLUSION The synergistic effects of BDSF and azoles may be useful as an alternative approach to control azole-resistant Candida infections.
Antifungal Agents ; pharmacology ; Burkholderia cenocepacia ; chemistry ; Candida albicans ; drug effects ; physiology ; Candidiasis ; drug therapy ; Drug Resistance, Fungal ; Fatty Acids, Monounsaturated ; adverse effects ; Fluconazole ; pharmacology ; Humans ; Microbial Sensitivity Tests ; Triazoles ; metabolism

Many of the fungicides and antibiotics currently available against plant pathogens are of limited use due to the emergence of resistant strains. In this study, we examined the effects of diphenyleneiodonium chloride (DPIC), an inhibitor of the superoxide producing enzyme NADPH oxidase, against fungal and bacterial plant pathogens. We found that DPIC inhibits fungal spore germination and bacterial cell proliferation. In addition, we demonstrated the potent antibacterial activity of DPIC using rice heads infected with the bacterial pathogen Burkholderia glumae which causes bacterial panicle blight (BPB). We found that treatment with DPIC reduced BPB when applied during the initial flowering stage of the rice heads. These results suggest that DPIC could serve as a new and useful antimicrobial agent in agriculture.
Agriculture ; Anti-Bacterial Agents ; Burkholderia ; Cell Proliferation ; Flowers ; Germination ; Head ; NADPH Oxidase ; Plants ; Spores, Fungal ; Superoxides