In this study, we investigated the temporal and spatial quantitative changes in the concentration of antibiotic resistance gene (ARG) markers in a municipal wastewater treatment plant (WWTP). Four ARGs conferring resistance to different classes of antibiotics ( ermB, sul 1, tet [W], and bla CTXM ) and a gene used as a proxy for ARG pollution ( intl1 ) were quantified in two separate sampling campaigns covering two and half years of operation of the WWTP. First, a systematic monthly monitoring of multiple points in the inlet and the outlet revealed an absolute decrease in the concentration of all analyzed ARGs. However, the relative abundance of sul 1 and intl 1 genes relative to the total bacterial load (estimated using the universal marker 16S rDNA) increased in the outlet samples as compared to the inlet. To pinpoint the exact stage of removal and/or enrichment within the WWTP, a second sampling including the stages of the biological treatment was performed bimonthly. This revealed a distinct enrichment of sul 1 and intl 1 genes during the biological treatment phase. Moreover, the temporal and spatial variations in ARG abundance patterns within the WWTP underscored the complexity of the dynamics associated with the removal of ARGs during wastewater treatment. Understanding these dynamics is pivotal for developing efficient strategies to mitigate the dissemination of ARGs in aquatic environments. Practitioner points Regular monitoring of ARG markers in WWTPs is essential to assess temporal and spatial changes, aiding in the development of effective mitigation strategies. Understanding the dynamics of ARG abundance during biological treatment is crucial for optimizing processes and minimizing dissemination in aquatic environments. Increased relative abundance of certain ARGs highlights potential enrichment during wastewater treatment, necessitating targeted interventions. Systematic monitoring of multiple points within WWTPs can provide valuable insights into the efficacy of treatment processes in reducing ARG levels over time. The complexity of ARG abundance patterns underscores the need to develop holistic approaches to tackle antibiotic resistance in wastewater systems.

Abstract In most habitats, fluctuating environmental conditions create periods of compromised survival for metabolically active organisms. In response, various survival strategies have evolved, including the formation of resilient resting cells. We assessed the biodiversity of the lysis‐resistant bacteria in three different environments by applying a harsh physicochemical treatment to the samples. The bacterial diversity of the lysis‐resistant fraction was compared with the bacterial diversity from the same environmental samples without the application of the enrichment method. As expected, in the lysis‐resistant fraction, a significantly higher relative abundance of endospore‐forming Firmicutes (for instance, Bacillus, Clostridium and Paenisporosarcina) was observed in comparison with the untreated samples. However, genera from which the existence of a resistant cell form is not yet reported were also highly enriched in comparison with the untreated samples. Our results suggest a more diversified repertoire of bacterial resistant cellular structures than previously thought.

Knowledge of associations between fungal hosts and their bacterial associates has steadily grown in recent years as the number and diversity of examinations have increased, but current knowledge is predominantly limited to a small number of fungal taxa and bacterial partners. Here, we screened for potential bacterial associates in over 700 phylogenetically diverse fungal isolates, representing 366 genera, or a tenfold increase compared with previously examined fungal genera, including isolates from several previously unexplored phyla. Both a 16 S rDNA-based exploration of fungal isolates from four distinct culture collections spanning North America, South America and Europe, and a bioinformatic screen for bacterial-specific sequences within fungal genome sequencing projects, revealed that a surprisingly diverse array of bacterial associates are frequently found in otherwise axenic fungal cultures. We demonstrate that bacterial associations with diverse fungal hosts appear to be the rule, rather than the exception, and deserve increased consideration in microbiome studies and in examinations of microbial interactions.

ABSTRACTBacteria–fungi interactions (BFIs) are essential in ecosystem functioning. These interactions are modulated not only by local nutritional conditions but also by the physicochemical constraints and 3D structure of the environmental niche. In soils, the unsaturated and complex nature of the substrate restricts the dispersal and activity of bacteria. Under unsaturated conditions, some bacteria engage with filamentous fungi in an interaction (fungal highways) in which they use fungal hyphae to disperse. Based on a previous experimental device to enrich pairs of organisms engaging in this interaction in soils, we present here the design and validation of a modified version of this sampling system constructed using additive printing. The 3D printed devices were tested using a novel application in which a target fungus, the common coprophilous fungus Coprinopsis cinerea, was used as bait to recruit and identify bacterial partners using its mycelium for dispersal. Bacteria of the genera Pseudomonas, Sphingobacterium and Stenotrophomonas were highly enriched in association with C. cinerea. Developing and producing these new easy-to-use tools to investigate how bacteria overcome dispersal limitations in cooperation with fungi is important to unravel the mechanisms by which BFIs affect processes at an ecosystem scale in soils and other unsaturated environments.

The production of a highly specialized cell structure called a spore is a remarkable example of a survival strategy displayed by bacteria in response to challenging environmental conditions. The detailed analysis and description of the process of sporulation in selected model organisms have generated a solid background to understand the cellular processes leading to the formation of this specialized cell. However, much less is known regarding the ecology of spore-formers. This research gap needs to be filled as the feature of resistance has important implications not only on the survival of spore-formers and their ecology, but also on the use of spores for environmental prospection and biotechnological applications.

The effect of three plant growth-promoting Bacillus strains inoculated either alone or as a consortium was tested on oat (Avena sativa) growth. The bioinoculants were applied as vegetative cells or endospores at low cell densities on the seeds and their effect was tested in sterile in vitro conditions, pot experiments, and a field trial. The in vitro seed germination assay showed that both individual bacterial inocula and bacterial consortia had positive effects on seed germination. Greenhouse pot experiments with sterile and non-sterile soil showed that consortia increased the total dry biomass of oat plants as compared to single strain inoculation and uninoculated controls. However, the positive impact on plant growth was less prominent when the bioinoculated strains had to compete with native soil microbes. Finally, the field experiment demonstrated that the consortium of vegetative cells was more efficient in promoting oat growth than the endospore consortium and the uninoculated control. Moreover, both consortia successfully colonized the roots and the rhizosphere of oat plants, without modifying the overall structure of the autochthonous soil microbial communities.

Lake Liambezi is located at the Eastern side of Caprivi Strip, straddling the border between Namibia and Botswana. The drainage basin of the lake is a large, flat wetland, including some woodlands, which contains a typically slow-flowing floodplain river (Seaman et al., 1978; Peel et al., 2015). The Lake changes its shape, size, and depth seasonally and over the years due to fluctuating contributions of water from its distinct source regions. The lake forms two elongated basins with a South-West to North-West direction joined by a main channel. The present study is based on multiple methods conducted on the watershed as well as on the lake. The aim is to give a paleoecological interpretation of the lake with isotopic and multi-proxy records. During the two field campaigns conducted during the dry season (September 2016) as well as at the end of the rainy season during March 2017, water from multiple sources of ground and surface waters were collected to better understand the drainage and hydrodynamics (Dyer, 2017). It was also established that the present climate supports a vegetation representing both the C3 and C4 photosynthetic cycles (Ballif, 2018). As they discriminate very differently against 13C during photosynthesis, such that the C-isotope composition of organic matter accumulated in soils or sediments may provide valuable information on the local ecosystem. In parallel to these geochemical studies three cores of 40 cm each were sampled in the lake. Sediments of the cores were first characterized by scanning electron microscopy that indicated a large fraction of diatoms in the sediments. Subsequent isotope analysis of carbon (13C/12C and 14C), nitrogen, hydrogen and oxygen of the organic matter and also RockEval measurements will evaluate the composition and quality of the organic matter. The lithogenic fraction is analysed via X-ray diffraction and fluorescence for the mineralogy and bulk chemical composition. Collectively, these analyses should allow for good estimates of the sedimentation rates, age of the sediment and a paleoecological interpretation that will then be compared to information obtained from endospore forming communities, a novel biological marker proposed in paleoecological reconstructions.

Aquatic ecosystems serve as a dissemination pathway and a reservoir of both antibiotic resistant bacteria (ARB) and antibiotic resistance genes (ARG). In this study, we investigate the role of the bacterial sporobiota to act as a vector for ARG dispersal in aquatic ecosystems. The sporobiota was operationally defined as the resilient fraction of the bacterial community withstanding a harsh extraction treatment eliminating the easily lysed fraction of the total bacterial community. The sporobiota has been identified as a critical component of the human microbiome, and therefore potentially a key element in the dissemination of ARG in human-impacted environments. A region of Lake Geneva in which the accumulation of ARG in the sediments has been previously linked to the deposition of treated wastewater was selected to investigate the dissemination of tet(W) and sul1, two genes conferring resistance to tetracycline and sulfonamide, respectively. Analysis of the abundance of these ARG within the sporobiome (collection of genes of the sporobiota) and correlation with community composition and environmental parameters demonstrated that ARG can spread across the environment with the sporobiota being the dispersal vector. A highly abundant OTU affiliated with the genus Clostridium was identified as a potential specific vector for the dissemination of tet(W), due to a strong correlation with tet(W) frequency (ARG copy numbers/ng DNA). The high dispersal rate, long-term survival, and potential reactivation of the sporobiota constitute a serious concern in terms of dissemination and persistence of ARG in the environment.

The introduction of antibiotics for both medical and non-medical purposes has had a positive effect on human welfare and agricultural output in the past century.However, there is also an important ecological legacy regarding the use of antibiotics and the consequences of increased levels of these compounds in the environment as a consequence of their use and disposal.This legacy was investigated by quantifying two antibiotic resistance genes (ARG) conferring resistance to tetracycline (tet(W)) and sulfonamide (sul1) in bacterial seed bank DNA in sediments.The industrial introduction of antibiotics caused an abrupt increase in the total abundance of tet(W) and a steady increase in sul1.The abrupt change in tet(W) corresponded to an increase in relative abundance from ca. 1960 that peaked around 1976.This pattern of accumulation was highly correlated with the abundance of specific members of the seed bank community belonging to the phylum Firmicutes.In contrast, the relative abundance of sul1 increased after 1976.This correlated with a taxonomically broad spectrum of bacteria, reflecting sul1 dissemination through horizontal gene transfer.The accumulation patterns of both ARGs correspond broadly to the temporal scale of medical antibiotic use.Our results show that the bacterial seed bank can be used to look back at the historical usage of antibiotics and resistance prevalence.

The introduction of antibiotics for both medical and non-medical purposes has had a positive effect on human welfare and agricultural output in the past century. However, there is also an important ecological legacy regarding the use of antibiotics and the consequences of increased levels of these compounds in the environment as a consequence of their use and disposal. This legacy was investigated by quantifying two antibiotic resistance genes (ARG) conferring resistance to tetracycline (tet(W)) and sulfonamide (sul1) in bacterial seed bank DNA in sediments. The industrial introduction of antibiotics caused an abrupt increase in the total abundance of tet(W) and a steady increase in sul1. The abrupt change in tet(W) corresponded to an increase in relative abundance from ca. 1960 that peaked around 1976. This pattern of accumulation was highly correlated with the abundance of specific members of the seed bank community belonging to the phylum Firmicutes. In contrast, the relative abundance of sul1 increased after 1976. This correlated with a taxonomically broad spectrum of bacteria, reflecting sul1 dissemination through horizontal gene transfer. The accumulation patterns of both ARGs correspond broadly to the temporal scale of medical antibiotic use. Our results show that the bacterial seed bank can be used to look back at the historical usage of antibiotics and resistance prevalence.

Endospore formation is a survival strategy found among some bacteria from the phylum Firmicutes. During endospore formation, these bacteria enter a morpho-physiological resting state that enhances survival under adverse environmental conditions. Even though endospore-forming Firmicutes are one of the most frequently enriched and isolated bacterial groups in culturing studies, they are often absent from diversity studies based on molecular methods. The resistance of the spore core is considered one of the factors limiting the recovery of DNA from endospores. We developed a method that takes advantage of the higher resistance of endospores to separate them from other cells in a complex microbial community using physical, enzymatic and chemical lysis methods. The endospore-only preparation thus obtained can be used for re-culturing or to perform downstream analysis such as tailored DNA extraction optimized for endospores and subsequent DNA sequencing. This method, applied to sediment samples, has allowed the enrichment of endospores and after sequencing, has revealed a large diversity of endospore-formers in freshwater lake sediments. We expect that the application of this method to other samples will yield a similar outcome.

The Saprolegnia-Achlya clade comprises species of major environmental and economic importance due to their negative impact on aquaculture and aquatic ecosystems by threatening fishes, amphibians, and crustaceans. However, their taxonomy and phylogenetic relationships remain unresolved and suffer from many inconsistencies, which is a major obstacle to the widespread application of molecular barcoding to identify pathogenic strains with quarantine implications. We assessed phylogenetic relationships of major genera using three commonly used markers (ITS, SSU rRNA, and LSU rRNA). A consensus tree of the three genes provided support for nine clades encompassing eleven documented genera and a new clade (SAP1) that has not been described morphologically. In the course of this study, we isolated a new species, Newbya dichotoma sp. nov., which provided the only culture available for this genus. In parallel, we attempted to summarize the evolution of traits in the different genera, but their successive reversals rendered the inference of ancestral states impossible. This highlights even more the importance of a bar-coding strategy for saprolegniacean parasite detection and monitoring.