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The degradation of freshwater systems by salt pollution is a threat to global freshwater resources. Salinization is commonly identified by increased specific conductance (conductivity), a proxy for salt concentrations. However, conductivity fails to account for the diversity of salts entering freshwaters and the potential implications this has on microbial communities and functions. We tested 4 types of salt pollution-MgCl 2 , MgSO 4 , NaCl, and Na 2 SO 4 -on bacterial taxonomic and functional α-, β-diversity of communities originating from streams in two distinct localities (Nebraska [NE] and Ohio [OH], USA). Community responses depended on the site of origin, with NE and OH exhibiting more pronounced decreases in community diversity in response to Na 2 SO 4 and MgCl 2 than other salt amendments. A closer examination of taxonomic and functional diversity metrics suggests that core features of communities are more resistant to induced salt stress and that marginal features at both a population and functional level are more likely to exhibit significant structural shifts based on salt specificity. The lack of uniformity in community response highlights the need to consider the compositional complexities of salinization to accurately identify the ecological consequences of instances of salt pollution., (© 2024 The Authors. Environmental Microbiology published by John Wiley & Sons Ltd.)

The factors that influence biodiversity and productivity of hydrothermal ecosystems are not well understood. Here we investigate the relationship between fluid mixing, biodiversity, and chemosynthetic primary productivity in three co-localized hot springs (RSW, RSN, and RSE) in Yellowstone National Park that have different geochemistry. All three springs are sourced by reduced hydrothermal fluid, but RSE and RSN receive input of vapour phase gas and oxidized groundwaters, with input of both being substantially higher in RSN. Metagenomic sequencing revealed that communities in RSN were more biodiverse than those of RSE and RSW in all dimensions evaluated. Microcosm activity assays indicate that rates of dissolved inorganic carbon (DIC) uptake were also higher in RSN than in RSE and RSW. Together, these results suggest that increased mixing of reduced volcanic fluid with oxidized fluids generates additional niche space capable of supporting increasingly biodiverse communities that are more productive. These results provide insight into the factors that generate and maintain chemosynthetic biodiversity in hydrothermal systems and that influence the distribution, abundance, and diversity of microbial life in communities supported by chemosynthesis. These factors may also extend to other ecosystems not supported by photosynthesis, including the vast subterranean biosphere and biospheres beneath ice sheets and glaciers., (© 2023 The Authors. Environmental Microbiology published by Applied Microbiology International and John Wiley & Sons Ltd.)

Heterotrophic bacteria hydrolyze high molecular weight (HMW) organic matter extracellularly prior to uptake, resulting in diffusive loss of hydrolysis products. An alternative 'selfish' uptake mechanism that minimises this loss has recently been found to be common in the ocean. We investigated how HMW organic matter addition affects these two processing mechanisms in surface and bottom waters at three stations in the North Atlantic Ocean. A pulse of HMW organic matter increased cell numbers, as well as the rate and spectrum of extracellular enzymatic activities at both depths. The effects on selfish uptake were more differentiated: in Gulf Stream surface waters and productive surface waters south of Newfoundland, selfish uptake of structurally simple polysaccharides increased upon HMW organic matter addition. The number of selfish bacteria taking up structurally complex polysaccharides, however, was largely unchanged. In contrast, in the oligotrophic North Atlantic gyre, despite high external hydrolysis rates, the number of selfish bacteria was unchanged, irrespective of polysaccharide structure. In deep bottom waters (> 4000 m), structurally complex substrates were processed only by selfish bacteria. Mechanisms of substrate processing-and the extent to which hydrolysis products are released to the external environment-depend on substrate structural complexity and the resident bacterial community., (© 2024 Applied Microbiology International and John Wiley & Sons Ltd.)

Aquatic bacteria frequently are divided into lifestyle categories oligotroph or copiotroph. Oligotrophs have proportionately fewer transcriptional regulatory genes than copiotrophs and are generally non-motile/chemotactic. We hypothesized that the absence of chemotaxis/motility in oligotrophs prevents them from occupying nutrient patches long enough to benefit from transcriptional regulation. We first confirmed that marine oligotrophs are generally reduced in genes for transcriptional regulation and motility/chemotaxis. Next, using a non-motile oligotroph (Ca. Pelagibacter st. HTCC7211), a motile copiotroph (Alteromonas macleodii st. HOT1A3), and [ 14 C]l-alanine, we confirmed that l-alanine catabolism is not transcriptionally regulated in HTCC7211 but is in HOT1A3. We then found that HOT1A3 took 2.5-4 min to initiate l-alanine oxidation at patch l-alanine concentrations, compared to <30 s for HTCC7211. By modelling cell trajectories, we predicted that, in most scenarios, non-motile cells spend <2 min in patches, compared to >4 min for chemotactic/motile cells. Thus, the time necessary for transcriptional regulation to initiate prevents transcriptional regulation from being beneficial for non-motile oligotrophs. This is supported by a mechanistic model we developed, which predicted that HTCC7211 cells with transcriptional regulation of l-alanine metabolism would produce 12% of their standing ATP stock upon encountering an l-alanine patch, compared to 880% in HTCC7211 cells without transcriptional regulation., (© 2023 The Authors. Environmental Microbiology published by Applied Microbiology International and John Wiley & Sons Ltd.)

For chronic wounds, biofilm infection is a critical issue because it can tip the scales toward an unhealing state. Biofilm-based wound therapy has been extensively advocated. However, point-of-care biofilm diagnosis still largely relies on clinical judgment. In this study, we aimed to develop a rapid tool for diagnosing wound biofilm presence by alcian blue staining. First, we sought to optimize alcian blue staining using a colorimetric-based approach to detect the biofilm, specifically targeting polysaccharides in the extracellular polymeric substances. Among examined transfer membranes and cationic detergents at various concentrations, we selected a positively charged nylon transfer membrane for sample loading, and 1% cetyl trimethyl ammonium chloride (CTAC) as the blocking solution. After sample loading and blocking, the membrane was immersed in alcian blue solution for staining, followed by immersion in 1% CTAC to decrease background noise. Each step required only 30 seconds, and the whole procedure was completed within a few minutes. In the second part of this study, we enrolled 31 patients with chronic wounds to investigate the predictive validity of biofilm detection for unhealed wounds at a 1-month follow-up visit. Among the 18 cases with positive wound biofilm staining, 15 wounds (83.3%) were not healed at the 1-month follow-up visit. Only three unhealed wounds (30%) produced in negative staining cases. This finding indicates that biofilm infection is associated with poor healing outcome for chronic wounds. Moreover, our staining results correlated well with the clinical microbiological culture assessment (83.9% consistency; 95.2% sensitivity, and 60% specificity). In conclusion, the modified alcian blue staining protocol used here represents a rapid and sensitive procedure for detecting biofilm in chronic wounds. This technique provides a practical point-of-care approach for detection of wound biofilm, the implementation of which may improve clinical outcomes for chronic wound patients. Additional studies are required to validate this method., (© 2020 The Wound Healing Society.)

Hydrogen (H 2 ) is enriched in hot springs and can support microbial primary production. Using a series of geochemical proxies, a model to describe variable H 2 concentrations in Yellowstone National Park (YNP) hot springs is presented. Interaction between water and crustal iron minerals yields H 2 that partition into the vapour phase during decompressional boiling of ascending hydrothermal fluids. Variable vapour input leads to differences in H 2 concentration among springs. Analysis of 50 metagenomes from a variety of YNP springs reveals that genes encoding oxidative hydrogenases are enriched in communities inhabiting springs sourced with vapour-phase gas. Three springs in the Smokejumper (SJ) area of YNP that are sourced with vapour-phase gas and with the most H 2 in YNP were examined to determine the fate of H 2 . SJ3 had the most H 2 , the most 16S rRNA gene templates and the greatest abundance of culturable hydrogenotrophic and autotrophic cells of the three springs. Metagenomics and transcriptomics of SJ3 reveal a diverse community comprised of abundant populations expressing genes involved in H 2 oxidation and carbon dioxide fixation. These observations suggest a link between geologic processes that generate and source H 2 to hot springs and the distribution of organisms that use H 2 to generate energy., (© 2019 Society for Applied Microbiology and John Wiley & Sons Ltd.)

Large amounts of antimicrobials are used in salmonid aquaculture in Chile. Most are used in marine aquaculture, but appreciable amounts are also employed in freshwater aquaculture. Much research and many publications have examined transferable antimicrobial resistance in bacteria isolated from marine salmon farms, but much less attention has been paid to this area in freshwater salmon farming. A recent paper by Domínguez et al. (2019) has as least in part remedied this situation. We now comment on some of its interpretations and have attempted to point out its areas of strength and weakness in light of the published scientific literature. Seen in this setting, the important results presented by Domínguez et al. (2019) underline the need for increased awareness of the challenge to animal and human health posed by excessive use of antimicrobials in aquaculture., (© 2019 Society for Applied Microbiology and John Wiley & Sons Ltd.)

In this invited paper, I give a personal description of my start as a young scientist studying the sulphur cycle of marine sediments. The initial objective was to quantify the processes experimentally using 35 S as a tracer. The results showed an unexpected large contribution of sulphate reduction to the oxidation of sediment organic matter. This implied that sulphate reducing bacteria can perform complete oxidation of their substrates to CO 2 . A quantification of sulphate reducing bacteria compared to measured rates of sulphate reduction revealed that the viable counting techniques available at the time vastly underestimated the true cell numbers. Through our introduction of microsensors for oxygen, sulphide and pH, the microbial oxidation of H 2 S could be analysed at high spatial resolution. Studies of the large, filamentous sulphur bacteria revealed surprising adaptations to sulphide oxidation based on anaerobic respiration with nitrate stored in vacuoles., (© 2019 Society for Applied Microbiology and John Wiley & Sons Ltd.)

The association between free-living amoebae and pathogenic bacteria is an issue that has gained great importance due to the environmental and health consequences that it implies. In this paper, we analyse the techniques to follow an epidemiological study to identify associations between genera, species, genotypes and subgenotypes of amoebae with pathogenic bacteria, analysing their evolution and considering their usefulness. In this sense, we highlight the combination of microscopic and molecular techniques as the most appropriate way to obtain fully reliable results as well as the need to achieve the standardization of these techniques to allow the comparison of both environmental and clinical results., (© 2013 Society for Applied Microbiology and John Wiley & Sons Ltd.)

The stoichiometry of prokaryotes (Bacteria and Archaea) can control benthic phosphorus (P) fluxes relative to carbon (C) and nitrogen (N) during organic matter remineralization. This paper presents the first experimental data on benthic microbial stoichiometry. We used X-ray microanalysis to determine C : N : P ratios of individual prokaryotes from C-limited Baltic Sea sediments incubated under oxic or anoxic conditions. At approximately 400:1, C : P ratios of prokaryotes from both oxic and anoxic incubations were higher than the Redfield ratio for marine organic matter (106:1), whereas prokaryotic C : N ratios (6.4:1) were close to the Redfield ratio. We conclude that high microbial C : P ratios contribute to the enhanced remineralization of P from organic matter relative to C and N observed in many low oxygen marine settings., (© 2012 Society for Applied Microbiology and Blackwell Publishing Ltd.)

Natural formation of organically bound chlorine is extensive in many environments. The enzymes associated with the formation of chlorinated organic matter are produced by a large variety of organisms. Little is known about the ecological role of the process, the key question being: why do microorganisms promote chlorination of organic matter? In a recent paper we discuss whether organic matter chlorination may be a result of antagonistic interactions among microorganisms. In the present paper we evaluate whether extracellular microbial formation of reactive chlorine may be used as a defence against oxygen stress, and we discuss whether this process is likely to contribute to the formation of chlorinated organic matter. Our analysis suggests that periodic exposure to elevated concentrations of reactive oxygen species is a common denominator among the multitude of organisms that are able to enzymatically catalyse formation of reactive chlorine. There is also some evidence suggesting that the production of such enzymes in algae and bacteria is induced by oxygen stress. The relative contribution from this process to the extensive formation of chlorinated organic matter in natural environments remains to be empirically assessed., (© 2012 Society for Applied Microbiology and Blackwell Publishing Ltd.)

In a mesotrophic reservoir, we examined the effects on the bacterioplankton of distinct consumers of bacteria, viruses and heterotrophic nanoflagellates, both alone and combined in an experiment using natural populations and in situ incubations in dialysis bags. Ribosomal RNA-targeted probes were employed as well as 16S RNA gene based PCR denaturing gradient gel electrophoresis (DGGE) to enumerate bacterial groups and assess bacterial community composition. We employed probes for Actinobacteria (HGC69a probe), Cytophaga-Flavobacterium-Bacteroidetes bacteria (CF319a probe), BET42a probe (Betaproteobacteria) and a subgroup-Betaproteobacteria (R-BT065 probe). We found consumer-specific effects on bacterial activity and diversity (against a background of CF and BET dominating all treatments) suggesting distinct vulnerabilities to the two sources of mortality. For example, growth rate of Actinobacteria was only positive in the presence of flagellates, while towards the end of the experiment (T(72-96 h)) growth rate of R-BT was only positive in the viruses only treatment. More specific data on how viruses and flagellates influenced Flectobacillus are shown in the companion paper. Highest richness (number of DGGE bands) was found in the virus only treatment and lowest when both consumers were present. In addition, we found suggestions of both antagonistic and synergistic interactions between the two sources of bacterial mortality. Notably, bactivory by flagellates was associated with reductions in bacterial diversity and increases in viral production.

The analysis of terminal restriction fragment length polymorphisms (T-RFLP) of 16S rRNA genes has proven to be a facile means to compare microbial communities and presumptively identify abundant members. The method provides data that can be used to compare different communities based on similarity or distance measures. Once communities have been clustered into groups, clone libraries can be prepared from sample(s) that are representative of each group in order to determine the phylogeny of the numerically abundant populations in a community. In this paper methods are introduced for the statistical analysis of T-RFLP data that include objective methods for (i) determining a baseline so that 'true' peaks in electropherograms can be identified; (ii) a means to compare electropherograms and bin fragments of similar size; (iii) clustering algorithms that can be used to identify communities that are similar to one another; and (iv) a means to select samples that are representative of a cluster that can be used to construct 16S rRNA gene clone libraries. The methods for data analysis were tested using simulated data with assumptions and parameters that corresponded to actual data. The simulation results demonstrated the usefulness of these methods in their ability to recover the true microbial community structure generated under the assumptions made. Software for implementing these methods is available at http://www.ibest.uidaho.edu/tools/trflp&#95;stats/index.php.

In some zones of Antarctica's cold and dry desert, the extinction of cryptoendolithic microorganisms leaves behind inorganic traces of microbial life. In this paper, we examine the transition from live microorganisms, through their decay, to microbial fossils using in situ microscopy (transmission electron microscopy, scanning electron microscopy in back-scattered electron mode) and microanalytical (energy dispersive X-ray spectroscopy) techniques. Our results demonstrate that, after their death, endolithic microorganisms inhabiting Commonwealth Glacier sandstone from the Antarctica McMurdo Dry Valleys become mineralized. In some cases, epicellular deposition of minerals and/or simply filling up of empty moulds by minerals leads to the formation of cell-shaped structures that may be considered biomarkers. The continuous deposition of allochthonous clay minerals and sulfate-rich salts fills the sandstone pores. This process can give rise to microbial fossils with distinguishable cell wall structures. Often, fossilized cell interiors were of a different chemical composition to the mineralized cell walls. We propose that the microbial fossil formation observed was induced by mineral precipitation resulting from inorganic processes occurring after the death of cryptoendolithic microorganisms. Nevertheless, it must have been the organic template that provoked the diffusion of mineral elements and gave rise to their characteristic distribution pattern inside the fossilized cells.

The exploitation of the metagenome for novel biocatalysts by functional screening is determined by the ability to express the respective genes in a surrogate host. The probability of recovering a certain gene thereby depends on its abundance in the environmental DNA used for library construction, the chosen insert size, the length of the target gene, and the presence of expression signals that are functional in the host organism. In this paper, we present a set of formulas that describe the chance of isolating a gene by random expression cloning, taking into account the three different modes of heterologous gene expression: independent expression, expression as a transcriptional fusion and expression as a translational fusion. Genes of the last category are shown to be virtually inaccessible by shotgun cloning because of the low frequency of functional constructs. To evaluate which part of the metagenome might in this way evade exploitation, 32 complete genome sequences of prokaryotic organisms were analysed for the presence of expression signals functional in E. coli hosts, using bioinformatics tools. Our study reveals significant differences in the predicted expression modes between distinct taxonomic groups of organisms and suggests that about 40% of the enzymatic activities may be readily recovered by random cloning in E. coli.

Ikaite tufa columns from the Ikka Fjord in south-western Greenland constitute a natural, stable environment at low temperature and with a pH ranging from neutral at the exterior to very alkaline (pH 10.4) at the interior of the column. Phylogenetic analysis of culturable organisms revealed ten different isolates representing three of the major bacterial divisions. Nine of the isolates showed 94-99% similarity to known sequences, whereas one isolate displayed a low degree of similarity (less than 90%) to a Cyclobacterium species. Seven of the isolates were shown to be cold active alkaliphiles, whereas three isolates showed optimal growth at neutral pH. Phylogenetic analysis of DNA isolated directly from the ikaite material demonstrated the presence of a microbial flora more diverse than the culturable isolates. Whereas approximately half of the phylotypes showed 90-99% similarity to known meso- or thermophilic alkaliphiles, the rest of the sequences displayed less than 90% similarity when compared to known 16S rRNA gene sequences in databases. Thus, in the present paper, we demonstrate that ikaite columns that host a specialized macroscopic flora and fauna also contain a unique, cold active, alkaliphilic microflora.

It is generally assumed that increased microbial diversity corresponds to increased catabolic potential and, hence, to better removal of metabolites and pollutants. Yet, microbial diversity, more specifically richness of species in environmental samples and sites, is difficult to assess. It is proposed to interpret this diversity more in the framework of Pareto's law, i.e. 20% of the species govern 80% of the energy flux of the ecosystem. Ecological studies should attempt to delineate the main energy fluxes and that group of species playing quantitative key roles in the system. Consequently, bioaugmentation should aim at the rearrangement of the group of organisms dominantly involved in the overall energy flux, so that specific catabolic traits necessary for the clean up of pollutants are part of that active group. For soil ecosystems, the capacity of plant roots as creators of physical and chemical discontinuity should be used more strategically to bring about such rearrangements. Overall, this paper identifies a number of ecological concepts, such as the Pareto law, the Gompertz model and plant community-induced microbial competence, which may, given careful underpinning, open new perspectives for microbial ecology and biodegradation.

The widespread use of methyl tert-butyl ether (MTBE) as a gasoline additive has resulted in a large number of cases of groundwater contamination. Bioremediation is often proposed as the most promising alternative after treatment. However, MTBE biodegradation appears to be quite different from the biodegradation of usual gasoline contaminants such as benzene, toluene, ethyl benzene and xylene (BTEX). In the present paper, the characteristics of a consortium degrading MTBE in liquid cultures are presented and discussed. MTBE degradation rate was fast and followed zero order kinetics when added at 100 mg l(-1). The residual MTBE concentration in batch degradation experiments ranged from below the detection limit (1 microg l(-1)) to 50 microg l(-1). The specific activity of the consortium ranged from 7 to 52 mgMTBE g(dw)(-1) h(-1) (i.e. 19-141 mgCOD g(dw) (-1) h(-1)). Radioisotope experiments showed that 79% of the carbon-MTBE was converted to carbon-carbon dioxide. The consortium was also capable of degrading a variety of hydrocarbons, including tert-butyl alcohol (TBA), tert-amyl methyl ether (TAME) and gasoline constituents such as benzene, toluene, ethylbenzene and xylene (BTEX). The consortium was also characterized by a very slow growth rate (0.1 d(-1)), a low overall biomass yield (0.11 gdw g(-1)MTBE; i.e. 0.040 gdw gCOD(-1)), a high affinity for MTBE and a low affinity for oxygen, which may be a reason for the slow or absence of MTBE biodegradation in situ. Still, the results presented here show promising perspectives for engineering the in situ bioremediation of MTBE.

In winter, free-living Capercaillies feed almost exclusively upon needles of various conifer species. An important part of the energy and protein requirements of captive Capercaillies, however, is supplied by food rich in starch and proteins. The fecal flora of captive Capercaillies is considerably different from the flora of free-living Capercaillies. Bacterial diseases are frequent in captivity. The aim of the present study is to determine whether the essential oils from conifer needles possess antibacterial properties against several bacteria, some of which are facultatively pathogenic for Capercaillies. Needles of Pinus sylvestris contain few tannins, which could be a reason for the Capercaillies' preference for these needles. The needles of Abies alba contain high amounts of tannins and essential oils, which could explain the refusal of Capercaillies to eat these needles. The antibacterial effect of commercially available essential oils on the bacteria is investigated with a filter paper disc method and in a liquid medium. The essential oils possess a bacteriostatic or a bactericidal effect against some of the tested bacteria. Feeding on natural forage plants may help prevent the colonization of the intestines by facultatively pathogenic bacteria. The authors discuss why Capercaillies, especially young birds that are to be released, should be fed with natural forage plants.