Assessing long-term ecological impacts of PCE contamination in groundwater using a flow cytometric fingerprint approach.

This study aims to develop and validate a comprehensive method for assessing ecological disturbances in groundwater ecosystems caused by tetrachloroethylene (PCE) contamination, utilizing flow cytometry (FCM) fingerprint approach. We hypothesized that the ecological disturbance resulting from PCE contamination would exhibit 'press disturbance', persisting over extended periods, and inducing notable phenotypic differences in the microbial community compared to undisturbed groundwater. We collected 40 groundwater samples from industrial district with a history of over twenty years of PCE contamination, along with 56 control groundwater from the national surveillance groundwater system. FCM revealed significant alterations in the phenotypic diversity of microbial communities in PCE-contaminated groundwater, particularly during the dry season. The presence of specific dechlorinating bacteria (Dehalococcoides, Dehalogenimonas, and Geobacter) and their syntrophic partners was identified as an indicator of contamination. Phenotypic diversity measures provided clearer and more direct reflections of contamination impact compared to taxonomic diversity measures. This study establishes FCM fingerprinting as a simple, robust, and accurate method for evaluating ecological disturbances, with potential applications in early warning systems and continuous monitoring of groundwater contamination. The findings not only underscore the sensitivity of FCM in detecting phenotypic variations induced by environmental stressors but also highlight its utility in understanding the complex dynamics of microbial communities in contaminated groundwater ecosystems.

Phenotypic convergence of bacterial adaption to sub-lethal antibiotic treatment.

Microorganisms can adapt quickly to changes in their environment, leading to various phenotypes. The dynamic for phenotypic plasticity caused by environmental variations has not yet been fully investigated. In this study, we analyzed the time-series of phenotypic changes in Staphylococcus cells during adaptive process to antibiotics stresses using flow cytometry and Raman spectroscopy. The nine antibiotics with four different mode of actions were treated in bacterial cells at a sub-lethal concentration to give adaptable stress. Although the growth rate initially varied depending on the type of antibiotic, most samples reached the maximum growth comparable to the control through the short-term adaptation after 24 h. The phenotypic diversity, which showed remarkable changes depending on antibiotic treatment, converged identical to the control over time. In addition, the phenotype with cellular biomolecules converted into a bacterial cell that enhance tolerance to antibiotic stress with increases in cytochrome and lipid. Our findings demonstrated that the convergence into the phenotypes that enhance antibiotic tolerance in a short period when treated with sub-lethal concentrations, and highlight the feasibility of phenotypic approaches in the advanced antibiotic treatment.

Transcriptional Response and Plant Growth Promoting Activity of Pseudomonas fluorescens DR397 under Drought Stress Conditions.

Drought is one of the most vulnerable factors that affect crop productivity. Little is known about plant-associated microbiomes and their functional roles in assisting plant growth under drought. We investigated the genetic and transcriptomic characteristics of opportunistic beneficial microorganisms that selectively alleviate stress through plant-bacteria interactions under drought. Pseudomonas fluorescens DR397 was isolated from the drought-prone rhizospheric soil of soybean and showed high metabolic activity at -1.25 Mpa. The genome of DR397 possesses several genes related to the synthesis of compatible solutes (choline and glycine-betaine), exopolysaccharides (alginate and cellulose), and secretion systems (type II, III, IV, and VI), as well as genes related to plant growth promotion (indole-3-acetic acid, transketolase, and thiamine phosphate synthesis). The expression of these genes was significantly upregulated (8- to 263-fold change) only under drought conditions with plant root exudate treatment, whereas subtle transcriptomic changes were observed under solely root exudate treatment. When DR397 was placed on both legume cultivars (Pisum sativum and Phaseolus vulgaris), growth was hardly affected under well-watered conditions, but the shoot and root growths were increased by up from 62.0% to 149.1% compared with the control group under drought conditions. These results provide fundamental insight on the plant-bacterial interactions that alleviate plant stress as an important ecological strategy for improving drought tolerance. IMPORTANCE Drought is a serious abiotic stress on plants as wells as the microbes that coexist with plants, which significantly lowers their fitness. The plant-bacterial interaction is an important strategy to enhance their fitness under drought. However, many knowledge gaps still exist in our understanding of transcriptomic features of bacteria interacting with plant under drought. Here, by investigating the transcriptomic profiles and pot cultivation with legume, we show that the interactions of Pseudomonas fluorescens DR397 with plants change with drought. We, therefore, provide a fundamental evidence of a hidden hero in the soil that promote plant fitness from external stress.

Raman-Deuterium Isotope Probing and Metagenomics Reveal the Drought Tolerance of the Soil Microbiome and Its Promotion of Plant Growth.

Drought has become a major agricultural threat leading crop yield loss. Although a few species of rhizobacteria have the ability to promote plant growth under drought, the drought tolerance of the soil microbiome and its relationship with the promotion of plant growth under drought are scarcely studied. This study aimed to develop a novel approach for assessing drought tolerance in agricultural land by quantitatively measuring microbial phenotypes using stable isotopes and Raman spectroscopy. Raman spectroscopy with deuterium isotope probing was used to identify the Raman signatures of drought effects from drought-tolerant bacteria. Counting drought-tolerant cells by applying these phenotypic properties to agricultural samples revealed that 0% to 52.2% of all measured single cells had drought-tolerant properties, depending on the soil sample. The proportions of drought-tolerant cells in each soil type showed similar tendencies to the numbers of revived pea plants cultivated under drought. The phenotype of the soil microbiome and plant behavior under drought conditions therefore appeared to be highly related. Studying metagenomics suggested that there was a reliable link between the phenotype and genotype of the soil microbiome that could explain mechanisms that promote plant growth in drought. In particular, the proportion of drought-tolerant cells was highly correlated with genes encoding phytohormone production, including tryptophan synthase and isopentenyl-diphosphate delta-isomerase; these enzymes are known to alleviate drought stress. Raman spectroscopy with deuterium isotope probing shows high potential as an alternative technology for quantitatively assessing drought tolerance through phenotypic analysis of the soil microbiome. IMPORTANCE Soil microbiome has played a critical role in the plant survival during drought. However, the drought tolerance of soil microbiome and its ability to promote plant growth under drought is still scarcely studied. In this study, we identified the Raman signature (i.e., phenotype) of drought effects from drought-tolerant bacteria in agricultural soil samples using Raman-deuterium isotope probing (Raman-DIP). Moreover, the number of drought-tolerant cells measured by Raman-DIP was highly related to the survival rate of plant cultivation under drought and the abundance of genes encoding phytohormone production alleviating drought stress in plant. These results suggest Raman-DIP is a promising technology for measuring drought tolerance of soil microbiome. This result give us important insight into further studies of a reliable link between phenotype and genotype of soil microbiome for future plant-bacteria interaction research.

Pollution level and reusability of the waste soil generated from demolition of a rural railway.

Railways are typically considered polluted from years of train operation. However, the pollution level of railway in a rural area, which is less exposed to hazardous material from trains and freights, is rarely assessed. This study evaluated common railway pollutants such as heavy metals, total petroleum hydrocarbons (TPHs) and polycyclic aromatic hydrocarbons (PAHs) and their chemical properties in the waste soil generated from the renovation of an old railway in rural area of Wonju, South Korea. Furthermore, lab-scale cultivation tests of peas (Pisum sativum) were performed to assess reusability of the waste soil as a soil amendment. Carbonaceous materials were found in the upper layer of the railway (0 to -40 cm) and the concentration of common railway pollutants was comparable to those of the agricultural land nearby. Specifically, total aromatic and aliphatic TPHs were below detection limit; and total PAHs < 1.0 mg kg-1 was 1000-times less than railway functional parts. Applying the carbonaceous waste soil improved the water holding capacity of soil by approximately 10% and sprouts formed on the soil with 10% waste soil composition had greater fresh weight, stem length, and root length than the control. Although this investigation was confined to a small length of the railway route, the results confirm environmental safety and the potential value of the waste generated from rural railways for the first time.

A comparison study of the potential risks induced in arable land and forest soils by carcass-derived pollutants.

Improper decisions concerning animal carcass disposal sites pose grave threats to environmental biosecurity. However, only a few studies have focused on the effects of different land-use types on the composition of carcass-derived pollutants and microbial responses to the disturbances. This study was conducted using soil microcosms with minced pork built from arable land and forest soils for 5 weeks. To compare the risk induced from different land-use types by carcass burial, the soil properties, the microbial community, and multiple-antibiotic-resistant bacteria were evaluated for microcosm containing 0, 1.5 and 7.5 g of minced pork. The abiotic properties, including pH, organic carbon, nitrogen and phosphorus compounds, significantly increased, regardless of the land-use types and applied load masses. The microbial diversity indices of the forest soil were reduced, whereas those of the arable land remained relatively stable. The disturbances produced from carcass-derived pollutants altered the bacterial community structures differently for the different land-use types. The treatment increased multiple-antibiotic-resistant bacteria in the both soil samples, although the increase in the forest soil was significantly less compared to the arable land soils.

Metagenomic SMRT Sequencing-Based Exploration of Novel Lignocellulose-Degrading Capability in Wood Detritus from Torreya nucifera in Bija Forest on Jeju Island.

Lignocellulose, composed mostly of cellulose, hemicellulose, and lignin generated through secondary growth of woody plant, is considered as promising resources for biofuel. In order to use lignocellulose as a biofuel, biodegradation besides high-cost chemical treatments were applied, but knowledge on the decomposition of lignocellulose occurring in a natural environment is insufficient. We analyzed the 16S rRNA gene and metagenome to understand how the lignocellulose is decomposed naturally in decayed Torreya nucifera (L) of Bija forest (Bijarim) in Gotjawal, an ecologically distinct environment. A total of 464,360 reads were obtained from 16S rRNA gene sequencing, representing diverse phyla; Proteobacteria (51%), Bacteroidetes (11%) and Actinobacteria (10%). The metagenome analysis using single molecules real-time sequencing revealed that the assembled contigs determined originated from Proteobacteria (58%) and Actinobacteria (10.3%). Carbohydrate Active enZYmes (CAZy)- and Protein families (Pfam)-based analysis showed that Proteobacteria was involved in degrading whole lignocellulose, and Actinobacteria played a role only in a part of hemicellulose degradation. Combining these results, it suggested that Proteobacteria and Actinobacteria had selective biodegradation potential for different lignocellulose substrates. Thus, it is considered that understanding of the systemic microbial degradation pathways may be a useful strategy for recycle of lignocellulosic biomass, and the microbial enzymes in Bija forest can be useful natural resources in industrial processes.

Short-term Effects of Great Cormorant Droppings on Water Quality and Microbial Community of an Artificial Agricultural Reservoir.

Agricultural reservoirs are established to improve the management of water resources. Waterbirds in protected waters have become a nuisance, however, as nutrients from fecal deposits transported by the waterbirds have served to severely deteriorate water quality. Despite the importance of clean water resources, the microecology of small agricultural reservoirs regularly colonized by transitory waterbirds are seldom reviewed. To improve our understanding of the influence of waterbirds on small bodies of water, a microcosm study was conducted using water and sediment from an agricultural reservoir inhabited by 300 to 500 great cormorants. Temporal changes in total nitrogen, total phosphorous, chemical oxygen demand, NH-N, PO-P, and chlorophyll-a concentrations, in addition to the microbial community, were evaluated for microcosms containing 0, 0.5, 1.0, and 5.0 g of feces collected from a great cormorant colony. Chemical analysis of the water microcosm revealed that all microcosms showed both immediate and prolonged increases in nutrients due to the addition of feces. Additionally, a mere 0.5 g of feces doubled the concentration of chlorophyll-a from 2.1 ± 0.99 to 5.2 ± 1.1 µg L within 1 mo. Nonmetric multidimensional scaling of the microbial community structure revealed disturbances in both water and sediment microcosms. Disturbances to the microbial community in the water microcosm were significant only when 5.0 g of feces was added; however, disturbances to sediment microbial communities were induced by a smaller mass of feces. These results confirm the short-term water quality impairment and shift in microbial community structure caused by waterbird droppings and bird colony surface runoff in an agricultural reservoir.