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Monitoring of stream water quality is a key element of water resource management worldwide, but methods that are commonly used in temperate habitats may not be appropriate in humid tropical systems. We assessed the influence of four land uses on microbial water quality in 21 streams in the Panama Canal Watershed over a one-year period, using a common culture-based fecal indicator test and 16S rDNA metabarcoding. Each stream was located within one of four land uses: mature forest, secondary forest, silvopasture, and traditional cattle pasture. Culturing detected total coliforms and Escherichia coli across all sites but found no significant differences in concentrations between land uses. However, 16S rDNA metabarcoding revealed variability in the abundance of coliforms across land uses and several genera that can cause false positives in culture-based tests. Our results indicate that culture-based fecal indicator bacteria tests targeting coliforms may be poor indicators of fecal contamination in Neotropical oligotrophic streams and suggest that tests targeting members of the Bacteroidales would provide a more reliable indication of fecal contamination.

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Studies on dinoflagellate cysts in the Arabian Sea (AS) are limited to the coastal waters, but no information from the deeper depths. The dinoflagellate cyst assemblages in surface sediment samples (0-2 cm) from the deeper depths (up to ~ 4500 m) of central (oxygen minimum zone (OMZ)) and southeastern (oligotrophic) AS revealed that the relatively good numbers of cyst concentrations reach deeper depths of OMZ (3505 m) and oligotrophic (4368 m) regions, but the former harbored more cyst concentrations than the latter. The cyst concentration and species count (including HAB species) recorded here are lower compared to the eastern (EAS) and western (WAS) AS, but the autotrophic cyst dominance (74-83%) at deeper depths is in contrast with the heterotrophic dominance in coastal AS. Of the recorded 41 cyst species (belonging to 18 genera), four species (cyst of Cladopyxis sp., Operculodinium janduchenei, Stelladinium bifurcatum, and Protoperidinium monospinum) from the deepest part of oligotrophic AS form the first report. In contrast, Spiniferites and Lingulodinium cysts were common occurrences. Taxonomic comparison with literature revealed (i) the prevalence of more cosmopolitan species (32 species) which could be due to the prevalence of large and small-scale lateral transport of cysts in oligotrophic regions followed by OMZ and coastal regions, respectively, and (ii) very few region-specific species, i.e., cyst of Protoperidinium latissimum, Lejeunecysta sabrina, cyst of Protoperidinium denticulatum in EAS and Impagidinium patulum, and I. strialatum, in WAS. Interestingly, variability in the morphometry was evident between the coastal and open oceans in some cosmopolitan cysts, e.g., Operculodinium centrocarpum and Lingulodinium machaerophorum. These findings from the less studied pelagic regions will contribute to the growing knowledge of dinoflagellate cyst distribution patterns and highlight the significance of cyst taxa and morphology as potential ecological indicators for AS.

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BACKGROUND: Oligotrophy and hypereutrophy represent the two extremes of lake trophic states, and understanding the distribution of bacterial communities across these contrasting conditions is crucial for advancing aquatic microbial research. Despite the significance of these extreme trophic states, bacterial community characteristics and co-occurrence patterns in such environments have been scarcely interpreted. To bridge this knowledge gap, we collected 60 water samples from Lake Fuxian (oligotrophic) and Lake Xingyun (hypereutrophic) during different hydrological periods. RESULTS: Employing 16S rRNA gene sequencing, our findings revealed distinct community structures and metabolic potentials in bacterial communities of hypereutrophic and oligotrophic lake ecosystems. The hypereutrophic ecosystem exhibited higher bacterial α- and ß-diversity compared to the oligotrophic ecosystem. Actinobacteria dominated the oligotrophic Lake Fuxian, while Cyanobacteria, Proteobacteria, and Bacteroidetes were more prevalent in the hypereutrophic Lake Xingyun. Functions associated with methanol oxidation, methylotrophy, fermentation, aromatic compound degradation, nitrogen/nitrate respiration, and nitrogen/nitrate denitrification were enriched in the oligotrophic lake, underscoring the vital role of bacteria in carbon and nitrogen cycling. In contrast, functions related to ureolysis, human pathogens, animal parasites or symbionts, and phototrophy were enriched in the hypereutrophic lake, highlighting human activity-related disturbances and potential pathogenic risks. Co-occurrence network analysis unveiled a more complex and stable bacterial network in the hypereutrophic lake compared to the oligotrophic lake. CONCLUSION: Our study provides insights into the intricate relationships between trophic states and bacterial community structure, emphasizing significant differences in diversity, community composition, and network characteristics between extreme states of oligotrophy and hypereutrophy. Additionally, it explores the nuanced responses of bacterial communities to environmental conditions in these two contrasting trophic states.

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17ß-Estradiol (E2) is a novel micro-pollutant that is widely distributed in aquatic sediments and has a universal toxicological effect on aquatic organisms. However, its ecological impact on aquatic microorganisms is not yet clear. In this study, we designed a simulation system for oligotrophic water deposition in the laboratory, analyzed the impact of different concentrations of E2 pollution on the carbon metabolism activity (carbon gas emission rate) of water microorganisms. Based on high-throughput sequencing results, we revealed the impact of E2 pollution on the community structure succession and metabolic function of bacteria, archaea, and methanogens in the simulated system, explored the impact mechanism of E2 pollution on microbial carbon metabolism in water bodies. Our results suggested that E2 significantly impacts the bacterial and archaeal community rather than the methanogen community, thereby indirectly inhibiting methane production. The achievements will bridge the theoretical gap between estrogen metabolism and carbon metabolism in sedimentary environments and contribute to enriching the ecological toxicology theory of steroid estrogen.

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The efficient nitrogen removal from micro-polluted source water is an international challenge to be solved urgently. However, the inner denitrification mechanism of native aerobic denitrifying bacterial communities in response to carbon scarcity remains relatively unclear. Here, the bacterial community XT6, screened from an oligotrophic reservoir, exhibited aerobic denitrifying capacity under low-carbon environments. Up to 76.79-81.64 % of total organic carbon (TOC) and 51.48-67.60 % of NO3--N were removed by XT6 within 48 h at C/N ratios of 2.0-3.0. Additionally, the nitrogen balance experiments further manifested that 26.27-38.13 % of NO3--N was lost in gaseous form. As the C/N ratio decreased, XT6 tended to generate more extracellular polymeric substances (EPS), with the tightly bound EPS showing the largest increase. Pseudomonas and Variovorax were quite abundant in XT6, constituting 59.69 % and 28.65 % of the total sequences, respectively. Furthermore, metagenomics analysis evidenced that XT6 removed TOC and nitrate mainly through the tricarboxylic acid cycle and aerobic denitrification. Overall, the abovementioned results provide a deeper understanding of the nitrogen metabolic pathways of indigenous aerobic denitrifying bacterial communities with low C/N ratios and offer useful guidance for controlling nitrogen pollution in oligotrophic ecosystems.

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Cyanobacteria can form complex interactions with heterotrophic microorganisms, but this relationship is susceptible to nutrient concentrations. Disentangling the cyanobacteria-bacteria interactions in relation to nutrient supply is essential to understanding their roles in geochemical cycles under global change. We hypothesize that enhanced nutrient supply in oligotrophic oceans can promote interactions among cyanobacteria and bacteria. Therefore, we investigated the planktonic bacteria and their interactions with cyanobacteria in relation to elevated nutrients caused by enhanced upwelling around a shallow and a deep seamount in the tropical western Pacific Ocean. We found obviously higher complexity of network occurred with significantly more cyanobacteria in the deep chlorophyll maximum layer of the shallow seamount when compared with that of the deep seamount. Cyanobacteria can shape bacterial interaction and community evenness in response to relatively high nutrient concentrations. The effects of the nutrients on cyanobacteria-related networks were further estimated based on the Tara Oceans data. Statistical analyses further showed a facilitative effect of nitrate concentrations on cyanobacteria-bacteria mutualistic interactions in the global oligotrophic ocean. By analysing the Tara Ocean macrogenomic data, we detected functional genes related to cyanobacteria-bacteria interactions in all samples, indicating the existence of a mutualistic relationship. Our results reveal cyanobacteria-bacteria interaction in response to nutrient elevation in oligotrophic ocean and highlight the potentially negative effects of global change on the bacterial community from the view of the bio-interaction.

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This is the first study providing long-term data on the dynamics of bees and wasps and their parasitoids for the evidence-based management of reed beds. Ten years ago, we identified Lipara (Chloropidae) - induced galls on common reed (Phragmites australis, Poaceae) as a critically important resource for specialized bees and wasps (Hymenoptera: Aculeata). We found that they were surprisingly common in relatively newly formed anthropogenic habitats, which elicited questions about the dynamics of bees and wasps and their parasitoids in newly formed reed beds of anthropogenic origin. Therefore, in the winter and spring of 2022/23, we sampled reed galls from the same set of reed beds of anthropogenic and natural origin as those in 2012/13. At 10 sites, the number of sampled galls was similar in both time periods (80-122% of the value from 2012/13); 12 sites experienced a moderate decline (30-79% of the value from 2012/13), and the number of galls at six sampling sites was only 3-23% of their abundance in 2012/13. Spontaneous development was associated with increasing populations. After 10 years of spontaneous development, the populations of bees and wasps (including their parasitoids) bound to Lipara-induced reed galls increased in abundance and species richness or remained at their previous levels, which was dependent on the sampling site. The only identified threat consisted of reclamation efforts. The effects of habitat age were limited, and the assemblages in habitats of near-natural and anthropogenic origin largely overlapped. However, several species were consistently present at lower abundances in the anthropogenic habitats and vice versa. In conclusion, we provided evidence-based support for the establishment of oligotrophic reed beds of anthropogenic origin as management tools providing sustainable habitats for specialized reed gall-associated aculeate hymenopteran inquilines, including the threatened species.

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The Wnt signaling pathway is a key molecular process during fracture repair. Although much of what we now know about the role of this pathway in bone is derived from in vitro and animal studies, the same cannot be said about humans. As such, we hypothesized that Wnt signaling will also be a key process in humans during physiological fracture healing as well as in the development of a nonunion (hypertrophic and oligotrophic). We further hypothesized that the expression of Wnt-signaling pathway genes/proteins would exhibit a differential expression pattern between physiological fracture callus and the pathological nonunion tissues. We tested these two hypotheses by examining the mRNA levels of key Wnt-signaling related genes: ligands (WNT4, WNT10a), receptors (FZD4, LRP5, LRP6), inhibitors (DKK1, SOST) and modulators (CTNNB1 and PORCN). RNA sequencing from calluses as well as from the two nonunion tissue types, revealed that all of these genes were expressed at about the same level in these three tissue types. Further, spatial expression experiments identified the cells responsible of producing these proteins. Robust expression was detected in osteoblasts for the majority of these genes except SOST which displayed low expression, but in contrast, was mostly detected in osteocytes. Many of these genes were also expressed by callus chondrocytes as well. Taken together, these results confirm that Wnt signaling is indeed active during both human physiological fracture healing as well as in pathological nonunions.

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Dissolved organic matter (DOM), the most active component in interstitial waters, determines the stability of heavy metals and secondary release in sediments. However, little is known about the composition and metal-binding patterns of DOM in interstitial water from oligotrophic lakes affected by different anthropogenic perturbations. Here, 18 interstitial water samples were prepared from sediments in agricultural, residential, tourist, and forest regions in an oligotrophic lake (Shengzhong Lake in Sichuan Province, China) watershed. Interstitial water quality and DOM composition, properties, and Cu(II)- and Pb(II)-binding characteristics were measured via physicochemical analysis, UV-vis spectroscopic, fluorescence excitation-emission matrix-parallel factor analysis (EEM-PARAFAC), and fluorescence titration methods. The DOM, which was produced mainly by microbial activities, had low molecular weights, humification degrees, and aromaticity. Based on EEM-PARAFAC results, the DOM was generally composed of tryptophan- (57.7%), terrestrial humic- (18.7%), microbial humic- (15.6%), and tyrosine-like (8.0%) substances. The DOM in the metal complexes was primarily composed of tryptophan-like substances, which accounted for ~42.6% of the DOM-Cu(II) complexes and ~72.0% of the DOM-Pb(II) complexes; however, microbial humic-like substances primarily contributed to the stability of DOM-Cu(II) (logKCu = 3.7-4.6) and DOM-Pb(II) (logKPb = 4.3-4.8). Water quality parameters did not significantly affect the stability of DOM-metal complexes. We demonstrated that the metal-binding patterns of DOM in interstitial water from oligotrophic lakes are highly dependent on microbial DOM composition and are affected by anthropogenic perturbations to a lesser extent.

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Oligotrophic deep-water lakes are unique and sensitive ecosystems with limited nutrient availability. Understanding bacterial communities within these lakes is crucial for assessing ecosystem health, biogeochemical cycling, and responses to environmental changes. In this study, we investigated the seasonal and vertical dynamics of both free-living (FL) and particle-attached (PA) bacteria in Lake Fuxian, a typical oligotrophic deep freshwater lake in southeast China. Our findings revealed distinct seasonal and vertical dynamics of FL and PA bacterial communities, driven by similar physiochemical environmental factors. PA bacteria exhibited higher α- and ß-diversity and were enriched with Proteobacteria, Cyanobacteria, Firmicutes, Patescibacteria, Planctomycetota, and Verrucomicrobiota, while FL bacteria were enriched with Actinobacteria and Bacteroidota. FL bacteria showed enrichment in putative functions related to chemoheterotrophy and aerobic anoxygenic photosynthesis, whereas the PA fraction was enriched with intracellular parasites (mainly contributed by Rickettsiales, Chlamydiales, and Legionellales) and nitrogen metabolism functions. Deterministic processes predominantly shaped the assembly of both FL and PA bacterial communities, with stochastic processes playing a greater role in the FL fraction. Network analysis revealed extensive species interactions, with a higher proportion of positively correlated edges in the PA network, indicating mutualistic or cooperative interactions. Cyanobium, Comamonadaceae, and Roseomonas were identified as keystone taxa in the PA network, underscoring potential cooperation between autotrophic and heterotrophic bacteria in organic particle microhabitats. Overall, the disparities in bacterial diversity, community composition, putative function, and network characteristics between FL and PA fractions highlight their adaptation to distinct ecological niches within these unique lake ecosystems.IMPORTANCEUnderstanding the diversity of microbial communities, their assembly mechanisms, and their responses to environmental changes is fundamental to the study of aquatic microbial ecology. Oligotrophic deep-water lakes are fragile ecosystems with limited nutrient resources, rendering them highly susceptible to environmental fluctuations. Examining different bacterial types within these lakes offers valuable insights into the intricate mechanisms governing community dynamics and adaptation strategies across various scales. In our investigation of oligotrophic deep freshwater Lake Fuxian in China, we explored the seasonal and vertical dynamics of two bacterial types: free-living (FL) and particle-attached (PA). Our findings unveiled distinct patterns in the diversity, composition, and putative functions of these bacteria, all shaped by environmental factors. Understanding these subtleties provides insight into bacterial interactions, thereby influencing the overall ecosystem functioning. Ultimately, our research illuminates the adaptation and roles of FL and PA bacteria within these unique lake environments, contributing significantly to our broader comprehension of ecosystem stability and health.

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1,4-Dioxane concentration in most contaminated water is much less than 1 mg/L, which cannot sustain the growth of most reported 1,4-dioxane-metabolizing pure cultures. These pure cultures were isolated following enrichment of mixed cultures at high concentrations (20 to 1,000 mg/L). This study is based on a different strategy: 1,4-dioxane-metabolizing mixed cultures were enriched by periodically spiking 1,4-dioxane at low concentrations (≤1 mg/L). Five 1,4-dioxane-metabolizing pure strains LCD6B, LCD6D, WC10G, WCD6H, and WD4H were isolated and characterized. The partial 16S rRNA gene sequencing showed that the five bacterial strains were related to Dokdonella sp. (98.3%), Acinetobacter sp. (99.0%), Afipia sp. (99.2%), Nitrobacter sp. (97.9%), and Pseudonocardia sp. (99.4%), respectively. Nitrobacter sp. WCD6H is the first reported 1,4-dioxane-metabolizing bacterium in the genus of Nitrobacter. The net specific growth rates of these five cultures are consistently higher than those reported in the literature at 1,4-dioxane concentrations <0.5 mg/L. Compared to the literature, our newly discovered strains have lower half-maximum-rate concentrations (1.8 to 8.2 mg-dioxane/L), lower maximum specific 1,4-dioxane utilization rates (0.24 to 0.47 mg-dioxane/(mg-protein ⋅ d)), higher biomass yields (0.29 to 0.38 mg-protein/mg-dioxane), and lower decay coefficients (0.01 to 0.02 d-1). These are characteristics of microorganisms living in oligotrophic environments.

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The issue of combined pollution in oligotrophic water has garnered increasing attention in recent years. To enhance the pollutant removal efficiency in oligotrophic water, the system containing Zoogloea sp. FY6 was constructed using polyester fiber wrapped sugarcane biochar and construction waste iron (PWSI), and the denitrification test of simulated water and actual oligotrophic water was carried out for 35 days. The experimental findings from the systems indicated that the removal efficiencies of nitrate (NO3--N), total nitrogen (TN), chemical oxygen demand (COD), and total phosphorus (TP) in simulated water were 88.61%, 85.23%, 94.28%, and 98.90%, respectively. The removal efficiencies of actual oligotrophic water were 83.06%, 81.39%, 81.66%, and 97.82%, respectively. Furthermore, the high-throughput sequencing data demonstrated that strain FY6 was successfully loaded onto the biological carrier. According to functional gene predictions derived from the Kyoto Encyclopedia of Genes and Genomes (KEGG) database, the introduction of PWSI enhanced intracellular iron cycling and nitrogen metabolism.

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The healing of bone fractures can become aberrant and lead to nonunions which in turn have a negative impact on patient health. Understanding why a bone fails to normally heal will enable us to make a positive impact in a patient's life. While we have a wealth of molecular data on rodent models of fracture repair, it is not the same with humans. As such, there is still a lack of information regarding the molecular differences between normal physiological repair and nonunions. This study was designed to address this gap in our molecular knowledge of the human repair process by comparing differentially expressed genes (DEGs) between physiological fracture callus and two different nonunion types, hypertrophic (HNU) and oligotrophic (ONU). RNA sequencing data revealed over ∼18,000 genes in each sample. Using the physiological callus as the control and the nonunion samples as the experimental groups, bioinformatic analyses identified 67 and 81 statistically significant DEGs for HNU and ONU, respectively. Out of the 67 DEGs for the HNU, 34 and 33 were up and down-regulated, respectively. Similarly, out of the 81 DEGs for the ONU, 48 and 33 were up and down-regulated, respectively. Additionally, we also identified common genes between the two nonunion samples; 8 (10.8 %) upregulated and 12 (22.2 %) downregulated. We further identified many biological processes, with several statistically significant ones. Some of these were related to muscle and were common between the two nonunion samples. This study represents the first comprehensive attempt to understand the global molecular events occurring in human nonunion biology. With further research, we can perhaps decipher new molecular pathways involved in aberrant healing of human bone fractures that can be therapeutically targeted.

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Introduction: The positive effect of intercropping on host plant growth through plant-soil feedback has been established. However, the mechanisms through which intercropping induces interspecific competition remain unclear. Methods: In this study, we selected young apple trees for intercropping with two companion plants: medium growth-potential Mentha haplocalyx Briq. (TM) and high growth-potential Ageratum conyzoides L. (TA) and conducted mixed intercropping treatment with both types (TMA) and a control treatment of monocropping apples (CT). Results: Our findings revealed that TM increased the under-ground biomass of apple trees and TA and TMA decreased the above-ground biomass of apple trees, with the lowest above-ground biomass of apple trees in TA. The above- and under-ground biomass of intercrops in TA and TMA were higher than those in TM, with the highest in TA, suggesting that the interspecific competition was the most pronounced in TA. TA had a detrimental effect on the photosynthesis ability and antioxidant capacity of apple leaves, resulting in a decrease in above-ground apple biomass. Furthermore, TA led to a reduction in organic acids, alcohols, carbohydrates, and hydrocarbons in the apple rhizosphere soil (FRS) compared to those in both soil bulk (BS) and aromatic plant rhizosphere soil (ARS). Notably, TA caused an increase in pentose content and a decrease in the hexose/pentose (C6/C5) ratio in FRS, while ARS exhibited higher hexose content and a higher C6/C5 ratio. The changes in exudates induced by TA favored an increase in taxon members of Actinobacteria while reducing Proteobacteria in FRS compared to that in ARS. This led to a higher eutrophic/oligotrophic bacteria ratio relative to TM. Discussion: This novel perspective sheds light on how interspecific competition, mediated by root exudates and microbial community feedback, influences plant growth and development.

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Neuston, situated at the air-sea interface, stands as a crucial frontier in the realm of the global warming. Despite its unique habitat, there remains a need to substantiate the composition, diel dynamic and biotic-abiotic interaction of neustonic zooplankton in the tropical seas. In this study, we present rare observational data on neustonic zooplankton (0-20 cm) in the oligotrophic tropical South China Sea (SCS) during the summer of 2022. A total of eighteen samples were collected and analyzed, revealing the presence of fourteen taxa from eight phyla. The most prevalent group was Cypridina, accounting for 33.7% of the total abundance, followed by copepods (29.0%) and jellyfish (10.9%). Within copepods, the genus Pontella exhibited the highest relative abundance (38.0%). Additionally, each neuston taxon displayed unique diel distribution patterns. Cypridina was the most abundant taxon during the night (40.4%), while it shifted to copepod dominance during the day (50.4%). Among copepods, genus Pontella and larvae were dominant groups at night (44.7%) and during the day (30.0%), respectively. Moreover, a multivariate biota-environment analysis demonstrated that temperature, pH, dissolved oxygen and Si(OH)4 significantly impacted neuston composition. Notably, both jellyfish and sea snails showed a significant positive correlation with temperature, suggesting their potential dominance in the neuston community in response to future global warming in the oligotrophic tropical seas. This study lays a robust foundation for recognizing the neuston community in the oceanic SCS, and helps evaluate the long-term risks to neuston habitats under climate changes.

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Deep sea is a vast, dark, and difficult-to-access terrain and is now looked upon as a unique niche harboring diverse microorganism. We used a metataxonomic approach to decipher the microbial diversity present in the water column (surface to near bottom), water overlaying the sediments, and the deep-sea sediments (up to 35 cm) from the Indian Contract Region (ICR) in the Central Indian Ocean Basin (CIOB). Samples were collected from #IRZ (Impact Reference Zone), #PRZ (Potential Reference Zone), and #BC20 (Control site, outside potential mining area) with an average water depth of 5,200 m. 16S rRNA (V3-V4 region) amplicon sequencing on the MiSeq platform resulted in 942,851 ASVs across 65 water and sediment samples. Higher prokaryotic diversity was observed below 200 m in the water column to the seafloor. Proteobacteria was the most dominant bacterial phylum among all the water samples while Firmicutes, Actinobacteria and, Bacteroidota dominated the sediments. Sediment (below 10 cm) was co-dominated by Firmicutes. Thermoplasmata was the dominant archaeal group in the water column while Crenarchaeota was in the sediments. BC20 was less diverse than IRZ and PRZ. Deep Sea microorganisms could play a vital role in the mineralization processes, nutrient cycling, and also different biogeochemical cycles.

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Environmental oligotrophic bacteria are suspected to be highly relevant carriers of antimicrobial resistance (AMR). However, there is a lack of validated methods for monitoring in the aquatic environment. Since extended-spectrum ß-lactamases (ESBLs) play a particularly important role in the clinical sector, a culturing method based on R2A-medium spiked with different combinations of ß-lactams was applied to quantify ß-lactamase-producing environmental bacteria from surface waters. In German surface water samples (n = 28), oligotrophic bacteria ranging from 4.0 × 103 to 1.7 × 104 CFU per 100 mL were detected on the nutrient-poor medium spiked with 3rd generation cephalosporins and carbapenems. These numbers were 3 log10 higher compared to ESBL-producing Enterobacteriales of clinical relevance from the same water samples. A MALDI-TOF MS identification of the isolates demonstrated, that the method leads to the isolation of environmentally relevant strains with Pseudomonas, Flavobacterium, and Janthinobacterium being predominant ß-lactam resistant genera. Subsequent micro-dilution antibiotic susceptibility tests (Micronaut-S test) confirmed the expression of ß-lactamases. The qPCR analysis of surface waters DNA extracts showed the presence of ß-lactamase genes (blaTEM, blaCMY-2, blaOXA-48, blaVIM-2, blaSHV, and blaNDM-1) at concentrations of 3.7 (±1.2) to 1.0 (±1.9) log10 gene copies per 100 mL. Overall, the results demonstrate a widespread distribution of cephalosporinase and carbapenemase enzymes in oligotrophic environmental bacteria that have to be considered as a reservoir of ARGs and contribute to the spread of antibiotic resistance.

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Matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) is routinely used as a rapid and cost-effective method for pathogen identification in clinical settings. In comparison, its performance in other microbiological fields, such as environmental microbiology, is still being tested, although isolates of environmental microbes are essential for in-depth in vivo studies of their biology, including biotechnological applications. We investigated the applicability of MALDI-TOF MS for the identification of bacterial isolates from a highly oligotrophic environment - Dinaric Karst caves, which likely harbor specific microorganisms. We cultured bacteria from the shell surface of the endemic mussel Congeria jalzici, one of the three known cave mussels in the world that lives in the Dinaric karst underground. The bacterial isolates were obtained by swabbing the shell surface of mussels living in microhabitats with different amounts of water: 10 air-exposed mussels, 10 submerged mussels, and 10 mussels in the hygropetric zone. A collection of 87 pure culture isolates was obtained, mostly belonging to the phylum Bacillota (72%), followed by Pseudomonadota (16%), Actinomycetota (11%), and Bacteroidota (1%). We compared the results of MALDI-TOF MS identification (Bruker databases DB-5989 and version 11, v11) with the results of 16S rDNA-based phylogenetic analysis, a standard procedure for bacterial identification. Identification to the genus level based on 16S rDNA was possible for all isolates and clearly outperformed the results from MALDI-TOF MS, although the updated MALDI-TOF MS database v11 gave better results than the DB-5989 version (85% versus 62%). However, identification to the species-level by 16S rDNA sequencing was achieved for only 17% of isolates, compared with 14% and 40% for the MALDI-TOF MS databases DB-5989 and v11 database, respectively. In conclusion, our results suggest that continued enrichment of MALDI-TOF MS libraries will result with this method soon becoming a rapid, accurate, and efficient tool for assessing the diversity of culturable bacteria from different environmental niches.

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The consumption of cycloalkanes is prevalent in low-temperature marine environments, likely influenced by psychrophilic microorganisms. Despite their significance, the primary active species responsible for marine cycloalkane degradation remain largely unidentified due to cultivation challenges. In this study, we provide compelling evidence indicating that the uncultured genus C1-B045 of Gammaproteobacteria is a pivotal participant in cycloalkane decomposition within China's marginal seas. Notably, the relative abundance of C1-B045 surged from 15.9% in the methylcyclohexane (MCH)-consuming starter culture to as high as 97.5% in MCH-utilizing extinction cultures following successive dilution-to-extinction and incubation cycles. We used stable isotope probing, Raman-activated gravity-driven encapsulation, and 16 S rRNA gene sequencing to link cycloalkane-metabolizing phenotype to genotype at the single-cell level. By annotating key enzymes (e.g., alkane monooxygenase, cyclohexanone monooxygenase, and 6-hexanolactone hydrolase) involved in MCH metabolism within C1-B045's representative metagenome-assembled genome, we developed a putative MCH degradation pathway.

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The nitrogen removal characteristics and microbial response of biochar-immobilized mixed aerobic denitrifying bacteria (BIADB) were investigated at 25 °C and 10 °C. BIADB removed 53.51 ± 1.72 % (25 °C) and 39.90 ± 4.28 % (10 °C) nitrate in synthetic oligotrophic water. Even with practical oligotrophic water, BIADB still effectively removed 47.66-53.21 % (25 °C), and 39.26-45.63 % (10 °C) nitrate. The addition of inorganic electron donors increased nitrate removal by approximately 20 % for synthetic and practical water. Bacterial and functional communities exhibited significant temperature and stage differences (P < 0.05), with temperature and total dissolved nitrogen being the main environmental factors. The dominant genera and keystone taxa exhibited significant differences at the two temperatures. Structural equation model analysis showed that dissolved organic matter had the highest direct and indirect effects on diversity and function, respectively. This study provides an innovative pathway for utilizing biochar and inorganic electron donors for nitrate removal from oligotrophic waters.

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