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 shifts induced by environmental pre-stressors modify antibiotic resistance in Staphylococcus aureus.

Introduction: Escalating prevalence of antibiotic resistance in Staphylococcus aureus has necessitated urgent exploration into the fundamental mechanisms underlying antibiotic resistance emergence, particularly in relation to its interaction with environmental stressors. This study aimed to investigate the effects of environmental stressors prior to antibiotic exposure on the antibiotic resistance of S. aureus. Methods: We used Raman spectroscopy and flow cytometry to measure prior stress-induced phenotypic alterations of S. aureus, and identified the association between phenotypic shifts and the antibiotic resistance. Results: The results revealed a multifaceted relationship between stressors and the development of antibiotic resistance. The stressors effectuate distinct phenotypic diversifications and subsequently amplify these phenotypic alterations following antibiotic treatments, contingent upon the specific mode of action; these phenotypic shifts in turn promote the development of antibiotic resistance in S. aureus. This study's findings demonstrated that the presence of pre-stress conditions triggered an augmentation of resistance to vancomycin (VAN), while concurrently attenuating resistance to norfloxacin. Marked shifts in Raman peaks associated with lipids and nucleic acids demonstrated correlations with elevated survival rates following VAN treatment. Conclusion: Consequently, these observations indicate that pre-stress conditions "prime" bacterial cells for differential responses to antibiotics and bear significant implications for formulating clinical therapeutic strategies.

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.

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.

Enhanced deflocculation of dehydrated sludge by rhamnolipid treatment coupled with thermal hydrolysis.

The effect of a biosurfactant, rhamnolipid (RL), coupled with a thermal treatment was investigated to determine its impact on improving the deflocculation of dehydrated sludge from wastewater treatment processes. Results showed that the RL treatment positively impacted sludge conditioning to weak acidity and hydrolyzed the carbohydrates and proteins released from the matrix of the extracellular polymeric substance. When RL was coupled with high temperature thermal treatments (65 and 95 °C), soluble chemical oxygen demand and chemical oxygen demand solubilization increased by 9.6-19.7 times and 13.4-29.3%, respectively. The RL treatment reduced antibiotic resistant bacteria by 5.4-98.4%, and antibiotic activity was further accelerated by high temperature thermal treatment. The combination of biosurfactant and thermal treatment can effectively deflocculate dehydrated sludge and should be considered an alternative technology for the sludge management process.

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.

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.