Oleispirillum naphthae gen. nov., sp. nov., a bacterium isolated from oil sludge, and proposal of Oleispirillaceae fam. nov.

A microaerophilic, Gram-negative, motile, and spiral-shaped bacterium, designated Y-M2T, was isolated from oil sludge of Shengli oil field. The optimal growth condition of strain Y-M2T was at 25 °C, pH 7.0, and in the absence of NaCl. The major polar lipid was phosphatidylethanolamine. The main cellular fatty acid was iso-C17  :  0 3-OH. It contained Q-9 and Q-10 as the predominant quinones. The DNA G+C content was 68.1 mol%. Strain Y-M2T showed the highest 16S rRNA gene sequence similarity to Telmatospirillum siberiense 26-4bT (91.1 %). Phylogenetic analyses based on 16S rRNA gene and genomes showed that strain Y-M2T formed a distinct cluster in the order Rhodospirillales. Genomic analysis showed that Y-M2T possesses a complete nitrogen-fixation cluster which is phylogenetically close to that of methanogene. The nif cluster, encompassing the nitrogenase genes, was found in every N2-fixing strain within the order Rhodospirillales. Phylogeny, phenotype, chemotaxonomy, and genomic results demonstrated that strain Y-M2T represents a novel species of a novel genus in a novel family Oleispirillaceae fam. nov. in the order Rhodospirillales, for which the name Oleispirillum naphthae gen. nov., sp. nov. was proposed. The type strain is Y-M2T (=CCAM 827T=JCM 34765T).

A natural fusion of flavodiiron, rubredoxin, and rubredoxin oxidoreductase domains is a self-sufficient water-forming oxidase of Trichomonas vaginalis.

Microaerophilic pathogens such as Giardia lamblia, Entamoeba histolytica, and Trichomonas vaginalis have robust oxygen consumption systems to detoxify oxygen and maintain intracellular redox balance. This oxygen consumption results from H2O-forming NADH oxidase (NOX) activity of two distinct flavin-containing systems: H2O-forming NOXes and multicomponent flavodiiron proteins (FDPs). Neither system is membrane bound, and both recycle NADH into oxidized NAD+ while simultaneously removing O2 from the local environment. However, little is known about the specific contributions of these systems in T. vaginalis. In this study, we use bioinformatics and biochemical analyses to show that T. vaginalis lacks a NOX-like enzyme and instead harbors three paralogous genes (FDPF1-3), each encoding a natural fusion product between the N-terminal FDP, central rubredoxin (Rb), and C-terminal NADH:Rb oxidoreductase domains. Unlike a "stand-alone" FDP that lacks Rb and oxidoreductase domains, this natural fusion protein with fully populated flavin redox centers directly accepts reducing equivalents of NADH to catalyze the four-electron reduction of oxygen to water within a single polypeptide with an extremely high turnover. Furthermore, using single-particle cryo-EM, we present structural insights into the spatial organization of the FDP core within this multidomain fusion protein. Together, these results contribute to our understanding of systems that allow protozoan parasites to maintain optimal redox balance and survive transient exposure to oxic conditions.

Microaerophilic Activated Sludge System for Ammonia Retention toward Recovery from High-Strength Nitrogenous Wastewater: Performance and Microbial Communities.

A transition to ammonia recovery from wastewater has started; however, a technology for sustainable nitrogen retention in the form of ammonia and organic carbon removal is still in development. This study validated a microaerophilic activated sludge (MAS) system to efficiently retain ammonia from high-strength nitrogenous wastewater. The MAS is based on conventional activated sludge (CAS) with aerobic and settling compartments. Low dissolved oxygen (DO) concentrations (<0.2 mg/L) and short solids retention times (SRTs) (<5 days) eliminated nitrifying bacteria. The two parallel MASs were successfully operated for 300 days and had ammonia retention of 101.7 ± 24.9% and organic carbon removal of 85.5 ± 8.9%. The MASs mitigated N2O emissions with an emission factor of <0.23%, much lower than the default value of CAS (1.6%). A short-term step-change test demonstrated that N2O indicated the initiation of nitrification and the completion of denitrification in the MAS. The parallel MASs had comparable microbial diversity, promoting organic carbon oxidation while inhibiting ammonia-oxidizing microorganisms (AOMs), as revealed by 16S rRNA gene amplicon sequencing, the quantitative polymerase chain reaction of functional genes, and fluorescence in situ hybridization of ß-proteobacteria AOB. The microbial analyses also uncovered that filamentous bacteria were positively correlated with effluent turbidity. Together, controlling DO and SRT achieved organic carbon removal and successful ammonia retention, mainly by suppressing AOM activity. This process represents a new nitrogen management paradigm.

Genomic insight into iron acquisition by sulfate-reducing bacteria in microaerophilic environments.

Historically, sulfate-reducing bacteria (SRB) have been considered to be strict anaerobes, but reports in the past couple of decades indicate that SRB tolerate exposure to O2 and can even grow in aerophilic environments. With the transition from anaerobic to microaerophilic conditions, the uptake of Fe(III) from the environment by SRB would become important. In evaluating the metabolic capability for the uptake of iron, the genomes of 26 SRB, representing eight families, were examined. All SRB reviewed carry genes (feoA and feoB) for the ferrous uptake system to transport Fe(II) across the plasma membrane into the cytoplasm. In addition, all of the SRB genomes examined have putative genes for a canonical ABC transporter that may transport ferric siderophore or ferric chelated species from the environment. Gram-negative SRB have additional machinery to import ferric siderophores and ferric chelated species since they have the TonB system that can work alongside any of the outer membrane porins annotated in the genome. Included in this review is the discussion that SRB may use the putative siderophore uptake system to import metals other than iron.

Shumkonia mesophila gen. nov., sp. nov., a novel representative of Shumkoniaceae fam. nov. and its potentials for extracellular polymeric substances formation and sulfur metabolism revealed by genomic analysis.

A microaerophilic, mesophilic, chemoorganoheterotrophic bacterium, designated Y-P2T, was isolated from oil sludge enrichment in China. Cells of the strain were Gram-stain-negative, non-motile, non-spore-forming, rod-shaped or slightly curved with 0.8-3.0 µm in length and 0.4-0.6 µm in diameter. The strain Y-P2T grew optimally at 25 °C (range from 15 to 30 °C) and pH 7.0 (range from pH 6.0 to 7.5) without NaCl. The major cellular fatty acids were C16:0, summed feature 3 (C16:1 ω7c and/or C16:1 ω6c), summed feature 8 (C18:1 ω7c and/or C18:1 ω6c). The main polar liquids of strain Y-P2T comprised phosphatidylethanolamine (PE) and phosphatidylglycerol (PG). The respiratory quinone was Q-10. Acetate and H2 were the fermentation products of glucose. The DNA G + C content was 66.0%. Strain Y-P2T shared the highest 16S rRNA gene sequence similarity (90.3-90.6%) with species within Oceanibaculum of family Thalassobaculaceae in Rhodospirillales. Phylogenetic analyses based on 16S rRNA gene sequences and genomes showed that strain Y-P2T formed a distinct evolutionary lineage within the order Rhodospirillales. On the basis of phenotypic, phylogenetic and phylogenomic data, we propose that strain Y-P2T represents a novel species in a novel genus, for which Shumkonia mesophila gen. nov., sp. nov., within a new family Shumkoniaceae fam. nov. The type strain is Y-P2T (= CCAM 826 T = JCM 34766 T).

Large Fractionation in Iron Isotopes Implicates Metabolic Pathways for Iron Cycling in Boreal Shield Lakes.

Stable Fe isotopes have only recently been measured in freshwater systems, mainly in meromictic lakes. Here we report the δ56Fe of dissolved, particulate, and sediment Fe in two small dimictic boreal shield headwater lakes: manipulated eutrophic Lake 227, with annual cyanobacterial blooms, and unmanipulated oligotrophic Lake 442. Within the lakes, the range in δ56Fe is large (ca. -0.9 to +1.8‰), spanning more than half the entire range of natural Earth surface samples. Two layers in the water column with distinctive δ56Fe of dissolved (dis) and particulate (spm) Fe were observed, despite differences in trophic states. In the epilimnia of both lakes, a large Δ56Fedis-spm fractionation of 0.4-1‰ between dissolved and particulate Fe was only observed during cyanobacterial blooms in Lake 227, possibly regulated by selective biological uptake of isotopically light Fe by cyanobacteria. In the anoxic layers in both lakes, upward flux from sediments dominates the dissolved Fe pool with an apparent Δ56Fedis-spm fractionation of -2.2 to -0.6‰. Large Δ56Fedis-spm and previously published metagenome sequence data suggest active Fe cycling processes in anoxic layers, such as microaerophilic Fe(II) oxidation or photoferrotrophy, could regulate biogeochemical cycling. Large fractionation of stable Fe isotopes in these lakes provides a potential tool to probe Fe cycling and the acquisition of Fe by cyanobacteria, with relevance for understanding biogeochemical cycling of Earth's early ferruginous oceans.

Parasulfuritortus cantonensis gen. nov., sp. nov., a microaerophilic sulfur-oxidizing bacterium isolated from freshwater sediment.

A novel sulfur-oxidizing bacterium, designated strain LSR1T, was enriched and isolated from a freshwater sediment sample collected from the Pearl River in Guangzhou, PR China. The strain was an obligate chemolithoautotroph, using thiosulfate or sulfide as an electron donor and energy source. Growth of strain LSR1T was observed at 15-40 °C, pH 6.0-7.5 and NaCl concentrations of 0-1.5 %. Strain LSR1T was microaerophilic, with growth only at oxygen content less than 10 %. Anaerobic growth was also observed when using nitrate as the sole electron acceptor. The major cellular fatty acids were C16 : 0 and summed feature 3 (comprising C16 : 1 ω7c and/or C16 : 1 ω6c). The DNA G+C content of the draft genome sequence was 67.5 mol%. Phylogenetic analysis based on 16S rRNA gene sequences indicated that strain LSR1T formed a lineage within the family Thiobacillaceae, showing sequence identities of 92.87, 92.33 and 90.80 % with its closest relative genera Sulfuritortus, Annwoodia and Thiobacillus, respectively. The genome of strain LSR1T contained multiple genes encoding sulfur-oxidizing enzymes that catalyse thiosulfate and sulfide oxidation, and the gene encoding cbb 3-type cytochrome c oxidase and bd-type quinol oxidase, which enables strain LSR1T to perform sulphur oxidation under microaerophilic conditions. On the basis of phenotypic, genotypic and phylogenetic results, strain LSR1T is considered to represent a novel species of a new genus Parasulfuritortus within the family Thiobacillaceae, for which the name Parasulfuritortus cantonensis gen. nov., sp. nov. is proposed. The type strain is LSR1T (=GDMCC 1.1549=JCM 33645).

Microaerophilic Oxidation of Fe(II) Coupled with Simultaneous Carbon Fixation and As(III) Oxidation and Sequestration in Karstic Paddy Soil.

Microaerophilic Fe(II)-oxidizing bacteria are often chemolithoautotrophs, and the Fe(III) (oxyhydr)oxides they form could immobilize arsenic (As). If such microbes are active in karstic paddy soils, their activity would help increase soil organic carbon and mitigate As contamination. We therefore used gel-stabilized gradient systems to cultivate microaerophilic Fe(II)-oxidizing bacteria from karstic paddy soil to investigate their capacity for Fe(II) oxidation, carbon fixation, and As sequestration. Stable isotope probing demonstrated the assimilation of inorganic carbon at a maximum rate of 8.02 mmol C m-2 d-1. Sequencing revealed that Bradyrhizobium, Cupriavidus, Hyphomicrobium, Kaistobacter, Mesorhizobium, Rhizobium, unclassified Phycisphaerales, and unclassified Opitutaceas were fixing carbon. Fe(II) oxidation produced Fe(III) (oxyhydr)oxides, which can absorb and/or coprecipitate As. Adding As(III) decreased the diversity of functional bacteria involved in carbon fixation, the relative abundance of predicted carbon fixation genes, and the amount of carbon fixed. Although the rate of Fe(II) oxidation was also lower in the presence of As(III), over 90% of the As(III) was sequestered after oxidation. The potential for microbially mediated As(III) oxidation was revealed by the presence of arsenite oxidase gene (aioA), denoting the potential of the Fe(II)-oxidizing and autotrophic microbial community to also oxidize As(III). Thisstudy demonstrates that carbon fixation coupled to Fe(II) oxidation can increase the carbon content in soils by microaerophilic Fe(II)-oxidizing bacteria, as well as accelerate As(III) oxidation and sequester it in association with Fe(III) (oxyhydr)oxides.

Campylobacter upsaliensis isolated from a giant hepatic cyst.

Campylobacter upsaliensis is an enteropathogenic bacterium in animals, and is also rarely isolated from humans, where it can cause enteritis and bacteremia. This report describes the first case of isolation of C. upsaliensis from an infected giant hepatic cyst. This bacterium could not be cultured from abscess punctuate in a usual Campylobacter-selection medium (charcoal cefoperazone deoxycholate agar medium), because of high concentration of cefoperazone as a selection agent. It could not identified by matrix-assisted laser desorption ionization-time of flight mass spectrum. Rather, it was identified as C. upsaliensis by whole genome sequencing, including by multilocus sequence typing.

A review on metronidazole: an old warhorse in antimicrobial chemotherapy.

The 5-nitroimidazole drug metronidazole has remained the drug of choice in the treatment of anaerobic infections, parasitic as well as bacterial, ever since its development in 1959. In contrast to most other antimicrobials, it has a pleiotropic mode of action and reacts with a large number of molecules. Importantly, metronidazole, which is strictly speaking a prodrug, needs to be reduced at its nitro group in order to become toxic. Reduction of metronidazole, however, only takes place under very low concentrations of oxygen, explaining why metronidazole is exclusively toxic to microaerophilic and anaerobic microorganisms. In general, resistance rates amongst the pathogens treated with metronidazole have remained low until the present day. Nevertheless, metronidazole resistance does occur, and for the treatment of some pathogens, especially Helicobacter pylori, metronidazole has become almost useless in some parts of the world. This review will give an account on the current status of research on metronidazole's mode of action, metronidazole resistance in eukaryotes and prokaryotes, and on other 5-nitroimidazoles in use.

Aliibacillus thermotolerans gen. nov., sp. nov.: a thermophilic and heterotrophic ammonia-oxidizing bacterium from compost.

A novel moderately thermophilic and heterotrophic ammonia-oxidizing bacterium, designated strain BM62T, was isolated from compost in the thermophilic stage in Harbin, China. Phylogenetic analysis based on the 16S rRNA gene indicated that strain BM62T belongs to the family Bacillaceae within the class Bacilli and was most closely related to Alteribacillus iranensis X5BT (only 94.6% sequence similarity). Cells of strain BM62T were Gram-positive, rod-shaped, motile by periflagella, catalase-positive and oxidase-negative. Growth of strain BM62T was observed at salinities of 0-4% (optimum 2-3%), temperatures of 35-65 °C (optimum 50 °C) and pH values of 5-9 (optimum pH 7). The major cellular fatty acid was iso-C16:0, and the predominant ubiquinone was MK-7. The peptidoglycan type is A1γ, and meso-diaminopimelic acid was the diagnostic diamino acid. The major polar lipids were diphosphatidylglycerol, phospholipid and phosphatidylglycerol. The G + C content of its genomic DNA was 36.5 mol%. Data from this polyphasic taxonomy study suggested that strain BM62T should be classified as the type strain of the type species of a new genus within the family Bacillaceae for which the name Aliibacillus thermotolerans gen. nov., sp. nov. is proposed. The type strain of the species Aliibacillus thermotolerans sp. nov. is BM62T (= DSM 101851T = CGMCC 1.15790T). The respective DPD Taxon Number is GA00057.

The role of dissolved oxygen content as a modulator of microbial polyhydroxyalkanoate synthesis.

Polyhydroxyalkanoates (PHAs) are a diverse class of bio-polymers synthesized by bacteria, usually during imbalanced growth conditions. Optimizing PHA productivity is highly dependent on the bioreactor oxygen transfer rate (OTR), which is an important consideration for process performance and economics, particularly with increasing scale. Relatively few in-depth studies are available regarding the effect of OTR and dissolved oxygen content (DOC) on PHA formation, synthesis rates, composition, and characteristics. This review examines past research studies on the effect of low DOC environments on production of short-chain length (scl-) PHAs, synthesized by both pure and mixed cultures, in order to identify opportunities and gaps concerning the effect of DOC on production of medium-chain length (mcl-) PHAs, an area that has not been studied in detail. The literature indicates that production of scl-PHA (a reductive process) acts as an electron sink allowing cells to maintain balanced redox state at low DOC. Conversely, production of mcl-PHA via fatty acid de novo synthesis (also a reductive process) does not occur to any significant extent in low DOC environments, while mcl-PHA synthesis from fatty acids (an oxidative process) can be promoted in low DOC environments. The monomer composition, molecular mass, as well as physical and thermal properties of the polymer can change in response to OTR, but further research in this area is required for both scl- and mcl-PHAs. Process design and management of bioreactor OTR in PHA production might therefore be directed by the final application of the polymer rather than cost considerations.

Innovative biocatalytic production of soil substrate from green waste compost as a sustainable peat substitute.

In the present work, a new simple and quick eco-friendly method is discussed to handle effectively the green wastes and produce a sustainable peat substitute of high quality on the large scale. Principal physicochemical parameters, i.e., temperature, moisture, specific weight, pH, electrical conductivity and, also, microorganisms, organic matter, humic substances, total Kjeldahl nitrogen and total organic carbon, C/N ratio, ash, metal content and phytotoxicity, were monitored systematically. Humic substances content values were interrelated to both C/N ratio and pH values and, similarly, bulk density, TOC, TKN, C/N, GI, ash and organic matter were found interconnected to each other. A novel biocatalyst, extremely rich in soil microorganisms, prepared from compost extracts and peaty lignite, accelerated the biotransformation. Zeolite was also employed. The compost does not demonstrate any phytotoxicity throughout the entire biotransformation process and has increased humic substances content. Both humic substances content and germination index can be employed as maturation indices of the compost. Addition of compost, processed for 60 days only, in cultivations of grass plants led to a significant increase in the stem mass and root size, annotating the significant contribution of the compost to both growth and germination. The product obtained is comparable to peat humus, useful as peat substitute and can be classified as a first class soil conditioner suitable for organic farming.

Anaerobic respiration: In vitro efficacy of Nitazoxanide against mitochondriate Acanthamoeba castellanii of the T4 genotype.

Acanthamoeba is an opportunistic protist pathogen that is responsible for serious human and animal infection. Being one of the most frequently isolated protists from the environment, it is likely that it readily encounters microaerophilic environments. For respiration under anaerobic or low oxygen conditions in several amitochondriate protists, decarboxylation of pyruvate is catalyzed by pyruvate ferredoxin oxidoreductase instead of pyruvate dehydrogenase. In support, Nitazoxanide, an inhibitor of pyruvate ferredoxin oxidoreductase, is effective and non-mutagenic clinically against a range of amitochondriate protists, Giardia intestinalis, Entamoeba histolytica and Trichomonas vaginalis. The overall aim of the present study was to determine in vitro efficacy of Nitazoxanide against Acanthamoeba castellanii. At micromolar concentrations, the findings revealed that Nitazoxanide neither affected A. castellanii growth or viability nor amoeba-mediated host cell monolayer damage in vitro or extracellular proteolytic activities. Similarly, microaerophilic conditions alone had no significant effects. In contrast, microaerophilic conditions together with Nitazoxanide showed amoebicidal effects and inhibited A. castellanii-mediated host cell monolayer damage as well as extracellular proteases. Using encystation assays, it was observed that Nitazoxanide inhibited trophozoite transformation into cysts both under aerophilic and microaerophilic conditions. Furthermore, pre-treatment of cysts with Nitazoxanide inhibited A. castellanii excystation. These findings are important in the identification of potential targets that could be useful against parasite-specific respiration as well as to understand the basic biology of the life cycle of Acanthamoeba.

Surface-Exposed Protein Moieties of Burkholderia cenocepacia J2315 in Microaerophilic and Aerobic Conditions.

Burkholderia cepacia complex infections remain life-threatening to cystic fibrosis patients, and due to the limited eradication efficiency of current treatments, novel antimicrobial therapies are urgently needed. Surface proteins are among the best targets to develop new therapeutic strategies since they are exposed to the host's immune system. A surface-shaving approach was performed using Burkholderia cenocepacia J2315 to quantitatively compare the relative abundance of surface-exposed proteins (SEPs) expressed by the bacterium when grown under aerobic and microaerophilic conditions. After trypsin incubation of live bacteria and identification of resulting peptides by liquid chromatography coupled with mass spectrometry, a total of 461 proteins with ≥2 unique peptides were identified. Bioinformatics analyses revealed a total of 53 proteins predicted as localized at the outer membrane (OM) or extracellularly (E). Additionally, 37 proteins were predicted as moonlight proteins with OM or E secondary localization. B-cell linear epitope bioinformatics analysis of the proteins predicted to be OM and E-localized revealed 71 SEP moieties with predicted immunogenic epitopes. The protegenicity higher scores of proteins BCAM2761, BCAS0104, BCAL0151, and BCAL0849 point out these proteins as the best antigens for vaccine development. Additionally, 10 of the OM proteins also presented a high probability of playing important roles in adhesion to host cells, making them potential targets for passive immunotherapeutic approaches. The immunoreactivity of three of the OM proteins identified was experimentally demonstrated using serum samples from cystic fibrosis patients, validating our strategy for identifying immunoreactive moieties from surface-exposed proteins of potential interest for future immunotherapies development.

Oxygen limitation favors the production of protein with antimicrobial activity in Pseudoalteromonas sp.

This study examined the effect of dissolved oxygen concentration on the production of biomass and metabolites with antimicrobial activity of Pseudoalteromonas sp cultured at 0, 150, 250, or 450 revolutions per minute (rev. min(-1)). Dissolved oxygen (D.O) was monitored during the fermentation process, biomass was quantified by dry weight, and antimicrobial activity was assessed using the disk diffusion method. The bacterium Pseudoalteromonas reached similar concentration of biomass under all experimental agitation conditions, whereas antimicrobial activity was detected at 0 and 150 rev. min(-1) registering 0% and 12% of D.O respectively corresponding to microaerophilic conditions. Antibiotic activity was severely diminished when D.O was above 20% of saturation; this corresponded to 250 or 450 rev. min(-1). SDS-PAGE electrophoresis revealed a protein with a molecular weight of approximately 80 kilodaltons (kDa) with antimicrobial activity. Pseudoalteromonas is capable of growing under oxic and microaerophilic conditions but the metabolites with antimicrobial activity are induced under microaerophilic conditions. The current opinion is that Pseudoalteromonas are aerobic organisms; we provide additional information on the amount of dissolved oxygen during the fermentation process and its effect on antimicrobial activity.

Metagenomic analysis of Fe(II)-oxidizing bacteria for Fe(III) mineral formation and carbon assimilation under microoxic conditions in paddy soil.

Microbially mediated Fe(II) oxidation is prevalent and thought to be central to many biogeochemical processes in paddy soils. However, we have limited insights into the Fe(II) oxidation process in paddy fields, considered the world's largest engineered wetland, where microoxic conditions are ubiquitous. In this study, microaerophilic Fe(II) oxidizing bacteria (FeOB) from paddy soil were enriched in gradient tubes with FeS, FeCO3, and Fe3(PO4)2 as iron sources to investigate their capacity for Fe(II) oxidation and carbon assimilation. Results showed that the highest rate of Fe(II) oxidation (k = 0.836 mM d-1) was obtained in the FeCO3 tubes, and cells grown in the Fe3(PO4)2 tubes yielded maximum assimilation amounts of 13C-NaHCO3 of 1.74% on Day 15. Amorphous Fe(III) oxides were found in all the cell bands with iron substrates as a result of microbial Fe(II) oxidation. Metagenomics analysis of the enriched microbes targeted genes encoding iron oxidase Cyc2, oxygen-reducing terminal oxidase, and ribulose-bisphosphate carboxylase, with results indicated that the potential Fe(II) oxidizers include nitrate-reducing FeOB (Dechloromonas and Thiobacillus), Curvibacter, and Magnetospirillum. By combining cultivation-dependent and metagenomic approaches, our results found a number of FeOB from paddy soil under microoxic conditions, which provide insight into the complex biogeochemical interactions of iron and carbon within paddy fields. The contribution of the FeOB to the element cycling in rice-growing regions deserves further investigation.

Pseudomonas aeruginosa Bacterioferritin Is Assembled from FtnA and BfrB Subunits with the Relative Proportions Dependent on the Environmental Oxygen Availability.

Ferritins are iron storage proteins assembled from 24 subunits into a spherical and hollow structure. The genomes of many bacteria harbor genes encoding two types of ferritin-like proteins, the bacterial ferritins (Ftn) and the bacterioferritins (Bfr), which bind heme. The genome of P. aeruginosa PAO1 (like the genomes of many bacteria) contains genes coding for two different types of ferritin-like molecules, ftnA (PA4235) and bfrB (PA3531). The reasons for requiring the presence of two distinct types of iron storage protein in bacterial cells have remained largely unexplained. Attempts to understand this issue in P. aeruginosa through the recombinant expression of the ftnA and bfrB genes in E. coli host cells, coupled to the biochemical and structural characterization of the recombinant 24-mer FtnA and 24-mer BfrB molecules, have shown that each of the recombinant molecules can form an Fe3+-mineral core. These observations led to the suggestion that 24-mer FtnA and 24-mer BfrB molecules coexist in P. aeruginosa cells where they share iron storage responsibilities. Herein, we demonstrate that P. aeruginosa utilizes a single heterooligomeric 24-mer Bfr assembled from FtnA and BfrB subunits. The relative content of the FtnA and BfrB subunits in Bfr depends on the O2 availability during cell culture, such that Bfr isolated from aerobically cultured P. aeruginosa is assembled from a majority of BfrB subunits. In contrast, when the cells are cultured in O2-limiting conditions, the proportion of FtnA subunits in the isolated Bfr increases significantly and can become the most abundant subunit. Despite the variability in the subunit composition of Bfr, the 24-mer assembly is consistently arranged from FtnA subunit dimers devoid of heme and BfrB subunit dimers each containing a heme molecule.

Zetaproteobacteria Pan-Genome Reveals Candidate Gene Cluster for Twisted Stalk Biosynthesis and Export.

Twisted stalks are morphologically unique bacterial extracellular organo-metallic structures containing Fe(III) oxyhydroxides that are produced by microaerophilic Fe(II)-oxidizers belonging to the Betaproteobacteria and Zetaproteobacteria. Understanding the underlying genetic and physiological mechanisms of stalk formation is of great interest based on their potential as novel biogenic nanomaterials and their relevance as putative biomarkers for microbial Fe(II) oxidation on ancient Earth. Despite the recognition of these special biominerals for over 150 years, the genetic foundation for the stalk phenotype has remained unresolved. Here we present a candidate gene cluster for the biosynthesis and secretion of the stalk organic matrix that we identified with a trait-based analyses of a pan-genome comprising 16 Zetaproteobacteria isolate genomes. The "stalk formation in Zetaproteobacteria" (sfz) cluster comprises six genes (sfz1-sfz6), of which sfz1 and sfz2 were predicted with functions in exopolysaccharide synthesis, regulation, and export, sfz4 and sfz6 with functions in cell wall synthesis manipulation and carbohydrate hydrolysis, and sfz3 and sfz5 with unknown functions. The stalk-forming Betaproteobacteria Ferriphaselus R-1 and OYT-1, as well as dread-forming Zetaproteobacteria Mariprofundus aestuarium CP-5 and Mariprofundus ferrinatatus CP-8 contain distant sfz gene homologs, whereas stalk-less Zetaproteobacteria and Betaproteobacteria lack the entire gene cluster. Our pan-genome analysis further revealed a significant enrichment of clusters of orthologous groups (COGs) across all Zetaproteobacteria isolate genomes that are associated with the regulation of a switch between sessile and motile growth controlled by the intracellular signaling molecule c-di-GMP. Potential interactions between stalk-former unique transcription factor genes, sfz genes, and c-di-GMP point toward a c-di-GMP regulated surface attachment function of stalks during sessile growth.

MALDI-TOF MS and 16S RNA Identification of Culturable Gastric Microbiota: Variability Associated with the Presence of Helicobacter pylori.

Helicobacter pylori is the main bacteria associated with gastroduodenal diseases. Recent studies have reported that gastric microbiota might be modified by the H. pylori colonization, favoring gastric lesions' development. In Chile, the region of La Araucanía concentrates a high risk of gastric cancer associated with Helicobacter pylori colonization, rurality, poverty, and Mapuche ethnicity. Hence, we aimed to identify the culturable gastric microbiota and characterize its variability at different stages of epithelial injury, based on its H. pylori colonization in dyspeptic patients from this Chilean region. Microaerophilic bacteria strains were isolated from antrum biopsies of 155 dyspeptic patients' biopsies and identified using MALDI-TOF MS or 16sRNA gene sequencing for non-pylori species identification, and UreC gene amplification for H. pylori confirmation. We found 48 species from 18 families, mainly belonging to Neisseriaceae (21.3%), Streptococcaceae (20.0%), Actynomicetaceae (9.0%), Enterobacteriaceae, and Lactobacillaceae (4.5%); however, Streptococcaceae and Actinomycetaceae families showed a significant reduction in samples infected with H. pylori, along with a considerably lower diversity of species. Our results revealed a microbiota modification due to H. pylori colonization associated with the gastric epithelial state, suggesting a potential microbiota role for developing and progressing gastric diseases.