Extracellular DNA (exDNA) is abundant in many habitats, including soil, sediments, oceans and freshwater as well as the intercellular milieu of metazoa. For a long time, its origin has been assumed to be mainly lysed cells. Nowadays, research is collecting evidence that exDNA is often secreted actively and is used to perform a number of tasks, thereby offering an attractive target or tool for biotechnological, medical, environmental and general microbiological applications. The present review gives an overview on the main research areas dealing with exDNA, depicts its inherent origins and functions and deduces the potential of existing and emerging exDNA-based applications. Furthermore, it provides an overview on existing extraction methods and indicates common pitfalls that should be avoided whilst working with exDNA.

Globally, inland waters emit over 2 Pg of carbon per year as carbon dioxide, of which the majority originates from streams and rivers. Despite the global significance of fluvial carbon dioxide emissions, little is known about their diel dynamics. Here we present a large-scale assessment of day- and night-time carbon dioxide fluxes at the water-air interface across 34 European streams. We directly measured fluxes four times between October 2016 and July 2017 using drifting chambers. Median fluxes are 1.4 and 2.1 mmol m⁻² h⁻¹ at midday and midnight, respectively, with night fluxes exceeding those during the day by 39%. We attribute diel carbon dioxide flux variability mainly to changes in the water partial pressure of carbon dioxide. However, no consistent drivers could be identified across sites. Our findings highlight widespread day-night changes in fluvial carbon dioxide fluxes and suggest that the time of day greatly influences measured carbon dioxide fluxes across European streams.

The analysis of environmental DNA (eDNA) is revolutionizing the monitoring of biodiversity as it allows to assess organismic diversity at large scale and unprecedented taxonomic detail. However, eDNA consists of an extracellular and intracellular fraction, each characterized by particular properties that determine the retrievable information on when and where organisms live or have been living. Here, we review the fractions of eDNA, describe how to obtain them from environmental samples and present a four‐scenario concept that aims at enhancing spatial and temporal resolution of eDNA‐based monitoring. Importantly, we highlight how the appropriate choice of eDNA fractions precludes misinterpretation of eDNA‐based biodiversity data. Finally, future avenues of research towards eDNA fraction‐specific analyses are outlined to unravel the full potential of eDNA‐based studies targeting micro‐ and macro‐organisms.

The ubiquity and relevance of extracellular DNA (exDNA) are well-known and increasingly gaining importance in many fields of application such as medicine and environmental microbiology. Although sources and types of exDNA are manifold, ratios of specific DNA-molecules inside and outside of living cells can give reliable information about the activity of entire systems and of specific microbial groups or species. Here, we introduce a method to discriminate between internal (iDNA), as well as bound and free exDNA, and evaluate various DNA fractions and related ratios (ex:iDNA) regarding their applicability to be used as a fast, convenient, and reliable alternative to more tedious RNA-based activity measurements. In order to deal with microbial consortia that can be regulated regarding their activity, we tested and evaluated the proposed method in comparison to sophisticated dehydrogenase- and RNA-based activity measurements with two anaerobic microbial consortia (anaerobic fungi and syntrophic archaea and a microbial rumen consortium) and three levels of resolution (overall activity, total bacteria, methanogenic archaea). Furthermore, we introduce a 28S rRNA gene-specific primer set and qPCR protocol, targeting anaerobic fungi (Neocallimastigomycota). Our findings show that the amount of actively released free exDNA (fDNA) strongly correlates with different activity measurements and is thus suggested to serve as a proxy for microbial activity.

Pesticide levels are generally monitored within agricultural areas, but are commonly not assessed at public places. To assess possible contamination of non-target areas, 71 public playgrounds located next to intensively managed apple and wine orchards were selected in four valleys of South Tyrol (northern Italy). Further, the impact of environmental site characteristics on the number and concentration of pesticides was assessed. Grass samples from the selected playgrounds were collected and screened for 315 pesticide residues using standard gas chromatography and mass spectrometry. Nearly half of the playgrounds (45%) were contaminated by at least one pesticide and a quarter (24%) by more than one. Eleven of the 12 different detected pesticides are classified as endocrine-active substances including the insecticide phosmet and the fungicide fluazinam showing the highest concentrations (0.069 and 0.26 mg kg−1, respectively). Additionally, one disinfectant and one preservation agent was found. Playgrounds in Venosta valley were most often contaminated (76% of all investigated playgrounds), highest concentrations were found in the Low Adige (2.02 mg kg−1). Pesticide concentrations were positively associated with areal proportion of apple orchards in the surroundings, the amount of rainfall and wind speed. In contrast, increasing global irradiance, opposite wind direction, increasing distance to agricultural sites and high wind speeds when pesticide application was not allowed were associated with decreasing pesticide contamination. This study is among the first investigating pesticide contamination of public playgrounds together with environmental factors in areas with pesticide-intensive agriculture at the beginning of the growing season. It is likely that playgrounds will be affected by more pesticides and higher concentrations over the course of the crop season. The result, that the majority of the detected pesticides are classified as endocrine active is worrisome as children are especially vulnerable. Hence, we recommend that pesticide risk assessments should better include protection measures for non-target areas.