Dynamic utilization of low-molecular-weight organic substrates across a microbial growth rate gradient

Constantly in flux, low-molecular-weight organic substances (LMWOSs) are at the nexus between microorganisms, plant roots, detritus, and the soil mineral matrix. Nominal oxidation state of carbon (NOSC) has been put forward as one way to parameterize microbial uptake rates of LMWOSs and efficiency of carbon incorporation into new biomass. In this study, we employed an ecophysiological approach to test these proposed relationships using targeted exometabolomics (1H-NMR, HR-LCMS) coupled with stable isotope (13C) probing. We assessed the role of compound class and oxidation state on uptake kinetics and substrate-specific carbon use efficiency (SUE) during the growth of three model soil microorganisms (Penicillium spinulosum, Paraburkholderia solitsugae, and Ralstonia pickettii) in media containing 34 common LMWOSs. Microbial isolates were chosen to span a gradient in growth rate (0.046-0.316 hr−1) and differ phylogenetically (a fungal isolate and two bacterial isolates). Clustered, co-utilization of LMWOSs occured for all three organisms, but temporal cluster separation was most apparent for P. solitsugae. Potential trends (p <0.05) for early utilization of more oxidized substrates were present for the two bacterial isolates (P. solitsugae and R. pickettii), but high variability (R2 > 0.15) and a small effect of NOSC indicate these are not useful relationships for prediction. The SUEs ranged from 0.16-0.99 and the hypothesized inverse relationship between NOSC and SUE was not observed. Thus, our results do not provide compelling support for NOSC as a predictive tool, implying that metabolic strategies of organisms may be more important than chemical identity in determining LMWOS cycling in soils.ImportanceCommunity-level observations from soils indicate that low-molecular-weight compounds of higher oxidation state tend to be depleted from soil solution faster and incorporated less efficiently into microbial biomass under oxic conditions. Here, we tested hypothetical relationships between substrate chemical characteristics and the order of substrate utilization by aerobic heterotrophs at the population-level in culture, using two bacterial isolates (Ralstonia pickettii and Paraburkholderia solitsugae) and one fungal isolate from soil (Penicillium spinulosum). We found weak relationships indicating earlier uptake of more oxidized substrates by the two bacterial isolates but no relationship for the fungal isolate. We found no relationship between substrate identity and substrate use efficiency. Our findings indicate that substrate chemical characteristics have limited utility for modeling the depletion of low-molecular-weight organics from soil solution and incorporation into biomass over broader phylogenetic gradients.

The Role of a Rapid Prevention of Ralstonia pickettii Growth during Dialysis in a Frail Patient

Ralstonia pickettii is an opportunistic bacillus found in Pseudomonas species, with the ability to induce systemic infections. We report the case of a 69-year-old man, with a clinical history of myeloma, Type IIdiabetes, renal failure (grade IV), and colon cancer, that developed a severe bacterial infection, with acute asthenia and a fever, that appeared at the end of dialysis. Using theMALDI-TOF technology, the bacillus Ralstonia pickettii was identified, and an antimicrobial treatment was quickly started with a rapid microbiological remission.

805. Outbreak of Ralstonia pickettii Bacteremia Caused by Contaminated Hydromorphone in a Universitary Hospital in Bogota, Colombia

Background Ralstonia pickettii are aerobic non fermenter gram negative bacilli isolated in water and soil. It is related to nosocomial infection outbreaks and considered an opportunistic pathogen. There have been outbreaks reports due to contaminated water systems and sterile drug solutions which mainly occurs during manufacturing. We present the report of an outbreak of R. pickettii bacteremia secondary to a contamination of hydromorphone vials. Methods In February 2021 an outbreak of R. pickettii bacteremia was identified. All isolates were from blood cultures with slow growth, thus indicating the culturing of liquid inputs, intravenous administration solutions and commonly used drugs among patients including hydromorphone. Mass spectrometry (MALDI-TOF) was used for the identification and automated microdilution to determine sensitivity to antimicrobials of the isolates and clonality analysis of genetic relationships was carried out using the DICE coefficient, UPGMA algorithm Results During the outbreak, 19 patients with R. pickettii bacteremia were identified The global attack rate was 1,9%. 11/19 (58%) were women and 13/19 (68%) of the isolations were from inward patients and 6/19 (32%) were from intensive care unit. Factors that could contribute to the appearance of the outbreak were underlying pathology, 2 patients with a diagnosis of diabetes mellitus, 10 patients with a diagnosis of arterial hypertension, 5 patients with obesity, 6 patients with heart disease, additionally 7 patients with a diagnosis of SARS COV 2 and 6 patients with the use of corticosteroids. The global attack rate was 1,9% and mortality was 31.5% (6 patients). R. pickettii was identified from two batches of hydromorphone by MALDI-TOF and the clonality study concluded that the isolates analyzed, were clonal with a 100% similarity. The associated mortality rate was 5/29 (26.3%). Conclusion We confirmed an outbreak of R. pickettii due to the contamination of two hydromorphone badges in Colombia. It is crucial to acknowledge the importance of infection control and surveillance during the COVID-19 pandemic as well as maintaining adequate quality control of medication production in order to avoid presenting this kind of outbreaks. Disclosures Sandra Liliana. Valderrama-Beltrán, MD, MSc, Biotoscana (Speaker’s Bureau)MSD (Grant/Research Support, Scientific Research Study Investigator, Research Grant or Support, Speaker’s Bureau)Pfiezer (Speaker’s Bureau)

The Biotransformation and Biodecolorization of Methylene Blue by Xenobiotic Bacterium Ralstonia pickettii

The biotransformation and biodecolorization of methylene blue (MB) dye using the bacterium Ralstonia pickettii was investigated. This experiment was conducted in a nutrient broth (NB) medium after adding MB at 100 mg L–1 concentration. Approximately 98.11% of MB was decolorized after 18 h of incubation. In addition, the metabolic products detected by LC-TOF/MS were Azure A (AA), thionine, leuco-MB, and glucose-MB, which indicated the MB degradation through a reductase that attacked the heterocyclic central chromophore group present in the structure. Moreover, azure A and thionine fragments resulted from the N-demethylase enzyme that attacked the auxochrome group. Thus, this research was assumed to be the first scientific report suggesting the potential to use R. pickettii in the biodecolorization and biotransformation of dye waste, particularly MB.

Iron-dependent mutualism between Chlorella sorokiniana and Ralstonia pickettii forms the basis for a sustainable bioremediation system

Mutualism between microalgae and bacteria is ubiquitous, but remains underexplored as a basis for biodegradation of anthropogenic pollutants. In industrial systems, poor iron uptake by microalgae limits growth, bioprocessing efficacy, and bioremediation potential. Iron supplementation is costly and ineffective because iron remains insoluble in aqueous medium and biologically unavailable. In aquatic environments, microalgae develop an association with bacteria that solubilize iron by production of siderophore, which increases the bioavailability of iron as a public good. Algae, in exchange, provides dissolved organic matter to bacteria to sustain such interkingdom associations. Therefore, using a case study of azo dye degradation, we combine environmental isolations and synthetic ecology as a workflow, establishing a microbial community to degrade industrially relevant Acid Black 1 dye. We create a mutualism between previously non-associated chlorophyte alga Chlorella sorokiniana and siderophore-producing bacterium Ralstonia pickettii, based on the eco-evolutionary principle of exchange of iron and carbon. This siderophore-mediated increased iron bioavailability increases reductive iron uptake, growth rate, and azoreductase-mediated dye degradation of microalga. In exchange, C. sorokiniana produces galactose, glucose, and mannose as major extracellular monosaccharides, supporting bacterial growth. We propose a mechanism whereby extracellular ferrireductase is crucial for azoreductase-mediated dye degradation in microalgae. Our work demonstrates that bioavailability of iron, which is often overlooked in industrial bio-designs, governs microalgal growth and enzymatic processes. Our results suggest that algal-bacterial consortia based on the active association are a self-sustainable mechanism to overcome existing challenges of micronutrient availability in bioremediation systems and their industrial translation.

P157 Microbiome Analysis Reveals That Ralstonia is Responsible for Decreased Renal Function in Patients with Ulcerative Colitis

Background Inflammatory bowel diseases (IBDs), such as ulcerative colitis (UC), are characterized by a disturbance of the normal gut microbiota and contribute to the development of chronic kidney disease (CKD). The incidence of CKD is higher in individuals with UC, but the causal link is unknown. Therefore, we investigated the role of gut microbiota in decreasing renal function in patients with UC Methods We performed 16S ribosomal DNA sequencing using ileocecal mucosal samples from nine individuals with UC and CKD (UC+CKD), 29 individuals with UC only, and 12 healthy controls. We also analyzed the operational taxonomic units, microbial diversity, and correlation with renal function. Co-culture assay using kidney organoids and Caco-2 cells was performed. Figure 1. The taxonomic composition of the gut microbiota of the study population and comparison of the 10 most abundant genera and species in the UC + CKD, UC, and control groups. Results Bacterial species diversity was significantly decreased in the UC+CKD group compared to that in the other groups based on Shannon and inverse Simpson indexes. At the genus level, Ralstonia had significantly greater abundance in the UC+CKD and UC groups compared to the control group. At the species level, unclassified Ralstonia species and Citrobacter portucalensis showed higher abundance in the UC+CKD group compared to the other groups. The relative abundance of Ralstonia showed a negative correlation with the estimated glomerular filtration rate (eGFR), but a positive correlation with the serum uric acid level. Ralstonia pickettii represented a negative correlation with eGFR, and induced damaging changes to kidney organoids in co-culture assay. Figure 2. The correlation between the relative abundance of Ralstonia with renal function (indicated by eGFR) and serum uric acid level. Effects of Ralstonia pickettii on Caco-2 cells and kidney organoids. (A) mRNA expression of pro-inflammatory cytokine genes in Caco-2 cells treated with and without Ralstonia pickettii, reflecting the inflammation seen in ulcerative colitis. Conclusion Gut microbial community profiles of the UC+CKD group are different from those of the UC group. Furthermore, Ralstonia pickettii contributes in decreasing renal function in patients with UC.

Streptococcus australis and Ralstonia pickettii as Major Microbiota in Mesotheliomas

The microbiota has been reported to be correlated with carcinogenesis and cancer progression. However, its involvement in the pathology of mesothelioma remains unknown. In this study, we aimed to identify mesothelioma-specific microbiota using resected or biopsied mesothelioma samples. Eight mesothelioma tissue samples were analyzed via polymerase chain reaction (PCR) amplification and 16S rRNA gene sequencing. The operational taxonomic units (OTUs) of the effective tags were analyzed in order to determine the taxon composition of each sample. For the three patients who underwent extra pleural pneumonectomy, normal peripheral lung tissues adjacent to the tumor were also included, and the same analysis was performed. In total, 61 OTUs were identified in the tumor and lung tissues, which were classified into 36 species. Streptococcus australis and Ralstonia pickettii were identified as abundant species in almost all tumor and lung samples. Streptococcus australis and Ralstonia pickettii were found to comprise mesothelioma-specific microbiota involved in tumor progression; thus, they could serve as targets for the prevention of mesothelioma.