Copper membrane monooxygenases (CuMMOs) oxidize ammonia, methane and some short-chain alkanes and alkenes. They are encoded by three genes, usually in an operon of
. We aligned
operons from 66 ...microbial genomes, including members of the
-,
-, and
and the candidate phylum NC10. Phylogenetic and compositional analyses were used to reconstruct the evolutionary history of the enzyme and detect potential lateral gene transfer (LGT) events. The phylogenetic analyses showed at least 10 clusters corresponding to a combination of substrate specificity and bacterial taxonomy, but with no overriding structure based on either function or taxonomy alone. Adaptation of the enzyme to preferentially oxidize either ammonia or methane has occurred more than once. Individual phylogenies of all three genes,
and
, closely matched, indicating that this operon evolved or was consistently transferred as a unit, with the possible exception of the methane monooxygenase operons in
, where the
gene has a distinct phylogeny from
and
. Compositional analyses indicated that some clusters of
operons (for example, the
in gammaproteobacterial methanotrophs and the
in betaproteobacterial nitrifiers) were compositionally very different from their genomes, possibly indicating recent lateral transfer of these operons. The combined phylogenetic and compositional analyses support the hypothesis that an ancestor of the nitrifying bacterium
was the donor of methane monooxygenase (pMMO) to both the alphaproteobacterial and gammaproteobacterial methanotrophs, but that before this event the gammaproteobacterial methanotrophs originally possessed another CuMMO (Pxm), which has since been lost in many species.
One of the most important micronutrients for bacterial growth is iron, whose bioavailability in soil is limited. Consequently, rhizospheric bacteria such as Pseudomonas fluorescens employ a range of ...mechanisms to acquire or compete for iron. We investigated the transcriptomic and proteomic effects of iron limitation on P. fluorescens Pf-5 by employing microarray and iTRAQ techniques, respectively. Analysis of this data revealed that genes encoding functions related to iron homeostasis, including pyoverdine and enantio-pyochelin biosynthesis, a number of TonB-dependent receptor systems, as well as some inner-membrane transporters, were significantly up-regulated in response to iron limitation. Transcription of a ribosomal protein L36-encoding gene was also highly up-regulated during iron limitation. Certain genes or proteins involved in biosynthesis of secondary metabolites such as 2,4-diacetylphloroglucinol (DAPG), orfamide A and pyrrolnitrin, as well as a chitinase, were over-expressed under iron-limited conditions. In contrast, we observed that expression of genes involved in hydrogen cyanide production and flagellar biosynthesis were down-regulated in an iron-depleted culture medium. Phenotypic tests revealed that Pf-5 had reduced swarming motility on semi-solid agar in response to iron limitation. Comparison of the transcriptomic data with the proteomic data suggested that iron acquisition is regulated at both the transcriptional and post-transcriptional levels.
Abstract
Human host-associated microbial communities in body sites can reflect health status based on the population distribution and specific microbial properties in the heterogeneous community. ...Bacteria identification at the single-cell level provides a reliable biomarker and pathological information for clinical diagnosis. Nevertheless, biosamples obtained from some body sites cannot offer sufficient sample volume and number of target cells as required by most of the existing single-cell isolation methods such as flow cytometry. Herein we report a novel integrated microfluidic system, which consists of a microemulsion module for single-bacteria encapsulation and a sequential microdroplet capture and release module for selectively extracting only the single-bacteria encapsulated in microdroplets. We optimize the system for a success rate of the single-cell extraction to be > 38%. We further verify applicability of the system with prepared cell mixtures (
Methylorubrum extorquens
AM1 and
Methylomicrobium album
BG8) and biosamples collected from human skin, to quantify the population distribution of multiple key species in a heterogeneous microbial community. Results indicate perfect viability of the single-cell extracts and compatibility with downstream analyses such as PCR. Together, this research demonstrates that the reported single-bacteria extraction system can be applied in microbiome and pathology research and clinical diagnosis as a clinical or point-of-care device.
(formerly
) AM1 is a methylotrophic bacterium with a versatile lifestyle. Various carbon sources including acetate, succinate and methanol are utilized by
AM1 with the latter being a promising ...inexpensive substrate for use in the biotechnology industry. Itaconic acid (ITA) is a high-value building block widely used in various industries. Given that no wildtype methylotrophic bacteria are able to utilize methanol to produce ITA, we tested the potential of
AM1 as an engineered host for this purpose. In this study, we successfully engineered
AM1 to express a heterologous codon-optimized gene encoding
aconitic acid decarboxylase. The engineered strain produced ITA using acetate, succinate and methanol as the carbon feedstock. The highest ITA titer in batch culture with methanol as the carbon source was 31.6 ± 5.5 mg/L, while the titer and productivity were 5.4 ± 0.2 mg/L and 0.056 ± 0.002 mg/L/h, respectively, in a scaled-up fed-batch bioreactor under 60% dissolved oxygen saturation. We attempted to enhance the carbon flux toward ITA production by impeding poly-β-hydroxybutyrate accumulation, which is used as carbon and energy storage, via mutation of the regulator gene
. Unexpectedly, ITA production by the
mutant strain was not higher even though poly-β-hydroxybutyrate concentration was lower. Genome-wide transcriptomic analysis revealed that
mutation in the ITA-producing strain led to complex rewiring of gene transcription, which might result in a reduced carbon flux toward ITA production. Besides poly-β-hydroxybutyrate metabolism, we found evidence that PhaR might regulate the transcription of many other genes including those encoding other regulatory proteins, methanol dehydrogenases, formate dehydrogenases, malate:quinone oxidoreductase, and those synthesizing pyrroloquinoline quinone and thiamine co-factors. Overall,
AM1 was successfully engineered to produce ITA using acetate, succinate and methanol as feedstock, further supporting this bacterium as a feasible host for use in the biotechnology industry. This study showed that PhaR could have a broader regulatory role than previously anticipated, and increased our knowledge of this regulator and its influence on the physiology of
AM1.
A Gram-negative, spiral-shaped, chemolithotrophic, ammonia-oxidizing bacterium, designated APG3 ᵀ, was isolated into pure culture from sandy lake sediment collected from Green Lake, Seattle, WA, USA. ...Phylogenetic analyses based on the 16S rRNA gene sequence showed that strain APG3 ᵀ belongs to cluster 0 of the genus Nitrosospira , which is presently not represented by described species, with Nitrosospira multiformis (cluster 3) as the closest species with a validly published name (identity of 98.6 % to the type strain). Strain APG3 ᵀ grew at 4 °C but could not grow at 35 °C, indicating that this bacterium is psychrotolerant. Remarkably, the strain was able to grow over a wide range of pH (pH 5–9), which was greater than the pH range of any studied ammonia-oxidizing bacteria in pure culture. The DNA G+C content of the APG3 ᵀ genome is 53.5 %, which is similar to that of Nitrosospira multiformis ATCC 25196 ᵀ (53.9 %) but higher than that of Nitrosomonas europaea ATCC 19718 (50.7 %) and Nitrosomonas eutropha C71 (48.5 %). The average nucleotide identity (ANI) calculated for the genomes of strain APG3 ᵀ and Nitrosospira multiformis ATCC 25196 ᵀ was 75.45 %, significantly lower than the value of 95 % ANI that corresponds to the 70 % species-level cut-off based on DNA–DNA hybridization. Overall polyphasic taxonomy study indicated that strain APG3 ᵀ represents a novel species in the genus Nitrosospira , for which the name Nitrosospira lacus sp. nov. is proposed (type strain APG3 ᵀ = NCIMB 14869 ᵀ = LMG 27536 ᵀ = ATCC BAA-2542 ᵀ).
Pseudomonas protegens Pf-5 (formerly Pseudomonas fluorescens) is a biocontrol bacterium that produces the siderophore enantio-pyochelin under conditions of iron starvation in a process that is often ...accompanied by the secretion of its biosynthesis intermediates, salicylic acid and dihydroaeruginoic acid. In this study, we investigated whether several transporters that are encoded by genes within or adjacent to the enantio-pyochelin biosynthetic cluster, serve as efflux systems for enantio-pyochelin and/or its intermediates. In addition, we determined whether these transporters have broad substrates range specificity using a Phenotype Microarray system. Intriguingly, knockouts of the pchH and fetF transporter genes resulted in mutant strains that secrete higher levels of enantio-pyochelin as well as its intermediates salicylic acid and dihydroaeruginoic acid. Analyses of these mutants did not indicate significant change in transcription of biosynthetic genes involved in enantio-pyochelin production. In contrast, the deletion mutant of PFL_3504 resulted in reduced transcription of the biosynthetic genes as well as decreased dihydroaeruginoic acid concentrations in the culture supernatant, which could either point to regulation of gene expression by the transporter or its role in dihydroaeruginoic acid transport. Disruption of each of the transporters resulted in altered stress and/or chemical resistance profile of Pf-5, which may reflect that these transporters could have specificity for rather a broad range of substrates.
An ammonia-oxidizing bacterium, strain D1FHS, was enriched into pure culture from a sediment sample retrieved in Jiaozhou Bay, a hyper-eutrophic semi-closed water body hosting the metropolitan area ...of Qingdao, China. Based on initial 16S rRNA gene sequence analysis, strain D1FHS was classified in the genus Nitrosococcus, family Chromatiaceae, order Chromatiales, class Gammaproteobacteria; the 16S rRNA gene sequence with highest level of identity to that of D1FHS was obtained from Nitrosococcus halophilus Nc4(T). The average nucleotide identity between the genomes of strain D1FHS and N. halophilus strain Nc4 is 89.5%. Known species in the genus Nitrosococcus are obligate aerobic chemolithotrophic ammonia-oxidizing bacteria adapted to and restricted to marine environments. The optimum growth (maximum nitrite production) conditions for D1FHS in a minimal salts medium are: 50 mM ammonium and 700 mM NaCl at pH of 7.5 to 8.0 and at 37°C in dark. Because pertinent conditions for other studied Nitrosococcus spp. are 100-200 mM ammonium and <700 mM NaCl at pH of 7.5 to 8.0 and at 28-32°C, D1FHS is physiologically distinct from other Nitrosococcus spp. in terms of substrate, salt, and thermal tolerance.
The ammonia-oxidizing obligate aerobic chemolithoautotrophic gammaproteobacterium,
, is omnipresent in the world's oceans and as such important to the global nitrogen cycle. We generated and compared ...high quality draft genome sequences of
strains isolated from the Northeast (AFC27) and Southeast (AFC132) Pacific Ocean and the coastal waters near Barbados at the interface between the Caribbean Sea and the North Atlantic Ocean (C-27) with the recently published Draft Genome Sequence of
Strain NS58 (West Pacific Ocean) and the complete genome sequence of
C-107, the type strain (ATCC 19707) isolated from the open North Atlantic, with the goal to identify indicators for the evolutionary origin of the species. The genomes of strains C-107, NS58, C-27, and AFC27 were highly conserved in content and synteny, and these four genomes contained one nearly sequence-identical plasmid. The genome of strain AFC132 revealed the presence of genetic inventory unknown from other marine ammonia-oxidizing bacteria such as genes encoding NiFe-hydrogenase and a non-ribosomal peptide synthetase (NRPS)-like siderophore biosynthesis module. Comparative genome analysis in context with the literature suggests that AFC132 represents a metabolically more diverse ancestral lineage to the other strains with C-107 and NS58 potentially being the youngest. The results suggest that the
species evolved by genome economization characterized by the loss of genes encoding catabolic diversity while acquiring a higher redundancy in inventory dedicated to nitrogen catabolism, both of which could have been facilitated by their rich complements of CRISPR/Cas and Restriction Modification systems.
Nitrification plays a crucial role in global nitrogen cycling and treatment processes. However, the relationships between the nitrifier guilds of ammonia-oxidizing bacteria (AOB) and ...nitrite-oxidizing bacteria (NOB) are still poorly understood, especially in freshwater habitats. This study examined the physiological interactions between the AOB and NOB present in a freshwater aquarium biofilter by culturing them, either together or separately, in a synthetic medium. Metagenomic and 16S rRNA gene sequencing revealed the presence and the draft genomes of
-like AOB as well as
-like NOB in the cultures, including the first draft genome of
. The nitrifiers exhibited different growth rates with different ammonium (NH
) or nitrite concentrations (50-1,500 μM) and the growth rates were elevated under a high bicarbonate (HCO
) concentration. The half-saturation constant (
for NH
), the maximum growth rate (μ
), and the lag duration indicated a strong dependence on the synergistic relationships between the two guilds. Overall, the ecophysiological and metagenomic results in this study provided insights into the phylogeny of the key nitrifying players in a freshwater biofilter and showed that interactions between the two nitrifying guilds in a microbial community enhanced nitrification.
Flexible and bio‐integrated electronics have attracted great attention due to their enormous contributions to personalized medical devices. Power sources, serving as one of the most important ...components, have been suffering from many problems, including deficient biocompatibility, poor stretchability, and unstable electrical outputs under deformed conditions, which limits the practical applications in flexible and bio‐integrated electronics. Here, we reported a fully stretchable magnesium (Mg)–air battery based on dual‐ions‐conducting hydrogels (SDICH). The high‐performance battery enables long‐term operation with lighting 120 lighting emitting diodes (LEDs) for over 5 h. Benefiting from the advanced materials and mechanical designs, the battery exhibits stability electrical outputs under stretching, which allows to operate ordinarily under various mechanical deformations without performance decay. Furthermore, the great biocompatibility of the battery offers great opportunity for biomedical applications, which is demonstrated by a self‐adaption wound dressing system. The in vitro and in vivo results prove that the self‐adaption wound dressing can effectively prevent wound inflammation and promote wound healing. By exploiting thermal feedback mechanics, the system can adjust antibiotic release rate and dosage spontaneously according to the real‐time wound conditions. The proposed fully stretchable Mg–air battery and self‐adaption wound dressing display great potential in skin‐integrated electronics and personalized medicine.
Stretchable Magnesium‐Air Battery Based on Dual Ions Conducting Hydrogel for Intelligent Biomedical Applications.
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