Applied and Environmental Microbiology最新文献

To develop countermeasures against phage-resistant bacteria without antibiotics, a detailed phenotypic characterization of phage-resistant Escherichia coli BW25113 was performed. Phage susceptibility testing of E. coli BW25113 deletion mutants involved in lipopolysaccharide (LPS) synthesis revealed that the first glucose residue of the LPS outer core was essential for infection by phage S127. From E. coli BW25113 cells that survived S127 exposure, four phage-resistant strains were isolated and characterized. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis showed that the phage-resistant strains had LPS with a smaller molecular mass compared with that of the E. coli BW25113 parental strain. Fluorescence microscopy after BacLight staining, along with comparisons of viable counts on non-selective versus selective media, indicated increased membrane permeability in the resistant strains, resulting in heightened sensitivity to monocaprin, a natural non-ionic surfactant. Furthermore, upon elevating the culture temperature from 30°C to 37°C, the resistant strains exhibited increased Congo Red binding and autoaggregation, which was not observed in the parental strain. Viability assays revealed that both the phage-resistant strains and deep rough mutants, such as ΔhldE and ΔwaaG, did not grow at 46°C. Notably, regrowth after phage S127 treatment at 37°C was significantly delayed in the E. coli BW25113 parental strain grown at 46°C than in that grown at 37°C. E. coli populations that became phage resistant owing to truncated LPS chains were highly sensitive to hydrophobic antibacterial substances and high temperatures, suggesting that these could be critical factors for controlling phage-resistant bacteria.

Importance: The application of phages in agriculture and food-producing environments often faces challenges in the control of phage-resistant bacteria. To effectively address this issue, a deeper understanding of the unique phenotypes associated with phage resistance is warranted. Few studies have suppressed the regrowth of phage-resistant populations without using antibiotics, based on detailed phenotypic characterization. Here, we report that the phage-resistant Escherichia coli population selected by lytic phage S127 was sensitive to elevated temperature and decreased viability at 46°C. Furthermore, Congo Red binding and autoaggregation, which have been reported to exhibit unique behaviors in E. coli deep rough mutants, were dependent on high culture temperature. Our findings highlight a novel, exploitable phenotype of phage resistance in host bacteria that could be applied to the biocontrol of phage resistance in foodborne pathogens without the use of antibiotics in practical settings.

Lignin degradation by white-rot fungi proceeds through oxidative depolymerization of lignin polymers, followed by metabolism of the resulting low-molecular-weight aromatic fragments. Among these, the catabolism of the hydroxyphenyl (H) unit of p-coumaric acid (p-CA) remains poorly understood in fungi. Here, we investigated the metabolism of p-CA by Trametes versicolor. Two group A flavoprotein monooxygenases (FPMOs), TvMNX3 and TvMNX4, catalyzed the hydroxylation of p-CA to caffeic acid. Recombinant expression and biochemical analyses revealed that TvMNX4 exhibited the highest catalytic efficiency toward both p-CA and 4-hydroxybenzoic acid, suggesting that it plays a major role in these hydroxylation reactions. Both enzymes also hydroxylated several p-CA derivatives, including 4-hydroxybenzaldehyde and 4-hydroxybenzyl alcohol, as well as other lignin-derived guaiacyl and syringyl compounds. Structural modeling and docking analyses indicated that the substrate-binding pocket-particularly residue Leu219 in TvMNX4-is critical for substrate accommodation and catalytic activity. Together, these findings suggest that T. versicolor degrades p-CA via hydroxylation mediated by group A FPMOs. To our knowledge, this is the first report identifying p-CA 3-hydroxylase activity in eukaryotic FPMOs, expanding our understanding of fungal aromatic catabolism.

Importance: White-rot fungi are key players in the global carbon cycle through lignin degradation, yet the intracellular pathways that catabolize lignin-derived aromatics remain largely unresolved. The hydroxyphenyl unit compound p-coumaric acid (p-CA) is a major lignin fragment, but the enzymes responsible for its conversion to caffeic acid (CFA) have not been previously identified in fungi. This study demonstrates that Trametes versicolor employs group A flavoprotein monooxygenases (FPMOs) TvMNX3 and TvMNX4 for the hydroxylation of p-CA and related metabolites, representing an unrecognized branch of the p-CA catabolic pathway. Beyond ecological significance, the capacity of TvMNX4 to generate bioactive phenolics such as CFA and piceatannol underscores its potential for biotechnological applications, including the sustainable synthesis of pharmaceuticals and polymer precursors.

Airborne bacteria represent a critical yet highly variable component of atmospheric ecosystems, driven by a complex interplay of local generation (LG) and regional dispersal (RD). Through extensive monitoring over 3 years (315 aerosol samples), we quantified airborne bacterial populations using quantitative PCR combined with high-throughput sequencing, revealing substantial fluctuations (2.8-5.8 log₁₀ copy·m^-3) characterized by dramatic spring peaks and summer minima. Strikingly, episodic dust events and PM₁₀ spikes from distant arid regions thousands of kilometers away increased local bacterial abundance by 5.5- and 6.2-fold (P < 0.05), respectively. Correlation network analysis identified strong positive associations among bacterial abundance, desert PM₁₀, and local PM₁₀, alongside negative relationships with temperature, humidity, and precipitation. Time-series analyses further revealed robust, synchronized annual cycles for bacterial abundance, desert PM₁₀, and local PM₁₀ (P < 0.05), with parametric modeling capturing a 4-week lag between desert dust emissions and subsequent local microbial peaks. Structural equation modeling provided quantitative confirmation that both LG and RD significantly influenced airborne bacterial dynamics (P < 0.05), with RD predominating during peak spring dust storm activity. Collectively, our findings highlight the substantial role of transcontinental dust transport as a primary source of airborne bacteria, complementing and sometimes overshadowing LGs in determining atmospheric bacterial communities.IMPORTANCEThis study enhances understanding of how airborne bacterial populations vary throughout the year in Busan, South Korea, by analyzing both local generation and long-distance transport. Over 3 years of continuous monitoring, we observed consistent spring peaks in bacterial abundance, closely linked to dust storms originating in the arid regions of China and Mongolia. These dust events transport large quantities of particles carrying bacteria over thousands of kilometers, temporarily raising local airborne bacterial levels above typical background levels. The findings show that while bacteria are continuously emitted from local sources, regional dust transport can be the dominant driver, particularly during the spring dust storm season. This combination of local and regional influences results in complex seasonal cycles of bacterial abundance. Understanding how these processes interact is critical for predicting changes in air quality, evaluating potential health risks, and recognizing the ecological connections that link distant desert environments with downwind areas.

Rhodococcus erythropolis NI86/21, isolated from maize rhizosphere in Hungary, possesses one of the largest genomes (8.046 Mb) within the species. The genome comprises a 6.83 Mb chromosome and 1.22 Mb of extrachromosomal elements, including three circular and two fragmented linear plasmids. Comparative analysis identified five horizontally acquired genomic islands (HGTi), totaling 0.64 Mb with mosaic-like architecture derived from plasmids, phages, and chromosomal segments of other Nocardiaceae. Liquid chromatography-tandem mass spectrometry (LC-MS/MS)-based proteomic analysis revealed a lower expression of genes located in HGT elements (53%) compared to core chromosomal genes (73%), indicating regulatory silencing of foreign DNA. Nevertheless, an inducible cytochrome P450 monooxygenase (CYP116) responsible for thiocarbamate and atrazine degradation is encoded on HGTi_V and actively expressed upon herbicide exposure. Strikingly, an identical CYP450 locus is present on a conjugative plasmid in Rhodococcus sp. TE1 isolated from thiocarbamate-treated soil in Canada, demonstrating independent acquisition of the same catabolic module from a high GC% content Rhodococcus, under similar selective pressure. Frequent recombination between chromosomal and mobile elements generates the observed mosaic-like HGT structures, which we found common for R. erythropolis strains. These results highlight extraordinary genomic plasticity and rapid adaptive evolution in Rhodococci, enabling efficient colonization of herbicide-contaminated agro-ecosystems.IMPORTANCERhodococcus erythropolis NI86/21 exemplifies how bacterial genomes evolve through horizontal gene transfer and mobile elements. Its unusually large, plastic genome contains extensive HGT islands and a high load of active transposons, which shape mosaic genomic architectures and hinder complete genome assembly. These horizontally acquired regions, although partially silenced, encode key adaptive functions such as an inducible CYP116 monooxygenase enabling thiocarbamate and atrazine degradation. Remarkably, an identical CYP116 module is present in Rhodococcus sp. TE1 from thiocarbamate-treated Canadian soil, demonstrating that similar environmental pressures can drive independent acquisition of the same biodegradation trait. Together, the dynamic transposon activity, mosaic HGT structure, and geographically convergent gene recruitment highlight the extraordinary genomic plasticity of R. erythropolis and underscore its rapid adaptive potential in agro-ecosystems, with implications for microbial evolution and bioremediation strategies.

Phosphorus forms and distribution in organic manures vary under different treatment conditions, thereby exerting distinct effects on the soil microbiome and soil phosphorus transformation process. This study examined the effects of a novel manure treated with hyper-thermophilic fermentation combined with Bacillus strain inoculation, compared with raw and composted manure, on the oat rhizosphere microbiome and phosphorus transformation across different soil types in a controlled pot experiment. Our findings demonstrate that hyper-thermophilic fermentation with Bacillus inoculation not only promotes the survival and abundance of the bacterial genus Bacillus but also selectively enriches the hyper-thermophilic bacterial genus Thermobifida in the fermented manure. Notably, the application of hyper-thermophilic fermented manure led to a significant enrichment of keystone species like Bacillus and Thermobifida across both soil types, relative to other manure applications. These genera emerged as key drivers of available phosphorus, phosphatase activity, and differential metabolites in the rhizosphere, exhibiting a synergistic effect on soil phosphorus transformation. Fermented manure exhibited superior performance relative to conventional composted manure, as it increased the phosphorus uptake rate of oats by 35.5% in black soil and 27.9% meadow soil, respectively, over a single growing season. Additionally, among all organic manures, the application of fermented manure significantly enhanced the sequestration of phosphorus from manure in the soils, with 78.0% in black soil and 56.9% in meadow soil. This consequently reduced P loss to 13.6% and 34.4% in the respective soil types.

Importance: Phosphate-solubilizing microorganisms (PSMs) are frequently proposed as catalysts for promoting phosphorus recycling; however, their performance is often inefficient or ineffective in the context of a circular bioeconomy within agricultural systems. This study introduces innovative concepts and methodologies by integrating hyper-thermophilic fermentation with heat-resistant phosphate-solubilizing Bacillus inoculation, thereby enhancing the effective phosphorus recovery and utilization from manure waste in sustainable agricultural practices.

A surge in sequenced myxobacteria catalyzed by advancements in long-read genome and metagenome sequencing has provided sufficient data to scrutinize the conserved biosynthetic gene clusters (BGCs) within the phylum Myxococcota. Provided the utility of myxobacteria in environmental nutrient cycles and discovery of novel therapeutic leads, we sought to determine any conserved specialized metabolism in the phylum. Using a pan-genome approach to analyze 11 genera and 195 sequenced genomes, including 10 newly reported myxobacterial isolates, we observed five conserved BGCs. All five clusters encode for characterized metabolites with established ecological roles for four of the metabolites, and none of the metabolites are known toxins. Validation of our approach was done by analyzing Myxococcota genera without sufficient sequenced representatives for pan-genome analysis to observe the presence/absence of these five clusters. This approach enabled observation of genus-level conservation of BGCs with varying degrees of confidence due to the diversity of sequenced species within each genus. The indigoidine BGC typically found in Streptomyces spp. was notably conserved in Melittangium; heterologous expression of the core biosynthetic gene bspA in Escherichia coli and subsequent detection of indigoidine confirmed the identity of the indigoidine cluster. Conserved BGCs in myxobacteria reveal maintenance of biosynthetic pathways and cognate metabolites with ecological roles as chemical signals and stress response; these observations suggest competitive specialization of secondary metabolism and toxin production in myxobacteria.

Importance: Critical contributions to soil nutrient cycles by predatory bacteria, including the Myxococcota, and utility as a resource for the discovery of novel enzymology and metabolism motive continued isolation and characterization of myxobacteria from the environment. Each of these motivating factors involves specialized metabolites produced by myxobacteria and the biosynthetic gene clusters (BGCs) responsible for their assembly. Primarily associated with the predatory lifestyles of myxobacteria, myxobacterial specialized metabolites have been pursued as therapeutic leads for novel antibacterials, antifungals, anthelmintics, and cancer therapies. Despite these efforts and the observation that nearly all genera within the Myxococcota have an extraordinary number of BGCs, there is no consensus view of the conserved BGCs in the phylum. Our study revealed the core BGCs consistently present throughout the phylum. By reporting these core specialized metabolites and their ecological roles, we hope to streamline the discovery and investigation of specialized metabolism in myxobacteria.