Five-Year Survey (2014 to 2018) of Azole Resistance in Environmental Aspergillus fumigatus Isolates from China.

A total of 191 soil samples from Hangzhou, China, were submitted to detect non-wild-type (non-WT) Aspergillus fumigatus and its associated mechanisms. There were 2 (4.7%), 13 (12.4%), and 31 (23.1%) isolates identified as non-WT in 2014, 2016, and 2018, respectively. The resistant mutations of TR34/L98H, TR46/Y121F/T289A, and TR34/L98H/S297T/F495I were found in 3, 5, and 5 non-WT isolates. The G448S mutation, previously only found in clinical settings, was detected in A. fumigatus from soil samples.

Concentration and Variety of Carbapenemase Producers in Recreational Coastal Waters Showing Distinct Levels of Pollution.

Carbapenemase-producing bacteria cause difficult-to-treat infections related to increased mortality in health care settings. Their occurrence has been reported in raw sewage, sewage-impacted rivers, and polluted coastal waters, which may indicate their spread to the community. We assessed the variety and concentration of carbapenemase producers in coastal waters with distinct pollution levels for 1 year. We describe various bacterial species producing distinct carbapenemases not only in unsuitable waters but also in waters considered suitable for primary contact.

Cross-resistance to fluconazole induced by exposure to the agricultural azole tetraconazole: an environmental resistance school?

This study aimed to investigate the influence of tetraconazole and malathion, both used in agricultural activities, on resistance to fluconazole, itraconazole and voriconazole in Candida parapsilosis ATCC 22019. The susceptibility to tetraconazole, malathion, fluconazole, itraconazole and voriconazole, through broth microdilution. Then, 12 independent replicates, were separated and exposed to four treatment groups, each one containing three replicates: G1: tetraconazole; G2: malathion; G3: fluconazole (positive control); G4: negative control. Replicates from G1, G2 and G3, were exposed to weekly increasing concentrations of tetraconazole, malathion and fluconazole, respectively, ranging from MIC/2 to 32 × MIC, throughout 7 weeks. The exposure to tetraconazole, but not malathion, decreased susceptibility to clinical azoles, especially fluconazole. The tetraconazole-induced fluconazole resistance is partially mediated by the increased activity of ATP-dependent efflux pumps, considering the increase in antifungal susceptibility after the addition of the efflux pump inhibitor, promethazine, and the increase in rhodamine 6G efflux and CDR gene expression in the G1 replicates. Moreover, MDR expression was only detected in G1 and G3 replicates, suggesting that MDR pumps are also involved in tetraconazole-induced fluconazole resistance. It is noteworthy that tetraconazole and fluconazole-treated replicates behaved similarly, therefore, resistance to azoles of clinical use may be a consequence of using azoles in farming activities.

Innovative wearable solutions: Semi-releasing ion-conductive lignin hydrogel sensors for enhanced practicability.

The severe negative effects of impurities adhering to the external surface of wearable devices can significantly influence the signal transmission, performance, and lifespan of hydrogel sensors. Herein, we developed an ion-conducting hydrogel sensor with a strong adhesive side and a non-adhesive side, similar to a "semi-releasing material." This hydrogel, formulated using deep eutectic solvents obtained from choline chloride and acrylic acid, contained lignin. This versatile material, exhibiting properties similar to semi-releasing materials, was treated with an AlCl3 solution on one side. Additionally, the hydrogel was successfully used as a highly adhesive strain sensor for real-time monitoring of various human activity signals. Moreover, the hydrogel demonstrated excellent environmental tolerance and conductivity. Lignin extracted from wood flour endowed the hydrogel sensor with excellent adhesion energy (up to 427.1 J/m2) and UV resistance. Treatment of hydrogels with AlCl3 completely eliminated their adhesiveness, thereby enhancing fracture elongation and tensile strength. This improvement can be attributed to the absence of carboxyl groups and the formation of a metal-phenolic network. The implementation of this convenient and efficient strategy provides a more feasible approach to address challenges related to impurity adhesion and signal transmission in flexible wearable devices.

Resilient green infrastructure: Navigating environmental resistance for sustainable development, social mobility in climate change policy.

This study explores the complex dynamics of environmental resistance, policy stability, skill development, and green initiatives in light of the growing worldwide concerns about climate change. COP27 and the G20 Summit show that as microcosm of the global discourse, provides a unique opportunity to study it. This research sought to thoroughly investigate, the links among green infrastructure, green technological innovation, policy stability, skill development, and their combined effects on social mobility and climate change (Case-1), it also discusses the global talk on climate change in COP27 and G20 (Case-2). In Case-1 data were analyzed through the use of structural equation modeling (SEM) by adopting a quantitative approach, and in Case-2 data were analyzed using theme analysis by applying a qualitative approach. Using a mixed-method research approach, the study surveyed 375 locals living close to the CPEC corridor quantitatively and interviewed ten important stakeholders, including elected officials, environmental activists, and community leaders, qualitatively. The survey highlighted the complex perspectives and experiences of citizens with green programs and environmental legislation in the CPEC zone. The study revealed the perceptions and experiences of residents regarding green initiatives and environmental policies within the CPEC region. Key stakeholders provided valuable insights into policy formulation and ongoing environmental sustainability efforts. The analysis unveiled intricate relationships between green infrastructure, technological innovation, policy stability, skill development, and their collective impact on climate change and social mobility. Notably, the study identified a critical research gap in understanding these dynamics within regions undergoing substantial economic development. Policy formulation and continuing environmental sustainability efforts were aided by key stakeholders' ideas. Green infrastructure, technological innovation, policy stability, skill development, and their overall influence on climate change and social mobility were all examined. Notably, the study found a critical research vacuum in understanding these processes inside rapidly developing economies. Policymakers, environmental groups, and communities managing the fine line between economic success and environmental responsibility will find great value in the findings. This study is unique because it examines issues on climate change from a local perspective in a region that is rapidly developing economically, it also adds value to the climate change challenges on the global level. This study presents a substantial theoretical contribution by examining the intricate interactions among environmental opposition, policy stability, skill development, and green initiatives within the CPEC against the backdrop of global climate change concerns.

Evaluation of the Effect of Mineral Oil Exposure on Changes in the Structure and Mechanical Properties of Polymer Parts Produced by Additive Manufacturing Techniques.

The paper describes the type of changes in the structure and mechanical properties of 3D printed shapes under the influence of mineral oil. The effects of a room (23 °C) and elevated temperature (70 °C) on 3D prints manufactured by the FDM method and stored in oil for 15, 30, and 60 days on the change of properties and structure were investigated. The samples were produced from ABS (poly(acrylonitrile-co-butadiene-co-styrene)), ASA (poly(acrylonitrile-co-styrene-co-acrylate), PLA (poly(lactic acid)), and HIPS (high-impact polystyrene). Tests related to the strength of the materials, such as the static tensile test and Charpy impact test, were carried out. The structure was evaluated using a scanning electron microscope, and changes in chemical structure were determined by conducting FTIR (Fourier transform infrared spectroscopy) and TGA (thermogravimetric analysis) tests. The analysis of the results provided important information about the impact of mineral oil on specific materials. This is critical for designing and manufacturing components that can withstand mineral oil exposure in real-world environments. The materials underwent varying changes. Strength increased for PLA by about 28%, remained unchanged for ABS and HIPS during exposure for 30 days, and decreased for ASA with extended exposure up to 14%.

Characterization of the Culturable Sporobiota of Spanish Olive Groves and Its Tolerance toward Environmental Challenges.

Olive agriculture presents an integral economic and social pillar of the Mediterranean region with 95% of the world's olive tree population concentrated in this area. A diverse ecosystem consisting of fungi, archaea, viruses, protozoa, and microbial communities-the soil microbiome-plays a central role in maintaining healthy soils while keeping up productivity. Spore-forming organisms (i.e., the sporobiota) have been identified as one of the predominant communities of the soil microbiome and are known for the wide variety of antimicrobial properties and extraordinary resistance. Hence, the aim of this work was to determine the culturable sporobiota of Spanish olive orchards and characterize its phenotypic properties toward common environmental challenges. A collection of 417 heat-resistant bacteria were isolated from five Spanish olive orchards. This collective was termed the "olive sporobiota." Rep-PCR clustering of representative isolates revealed that they all belonged to the group of Bacillus spp., or closely related species, showing a great variety of species and strains. Representative isolates showed susceptibility to common antibiotics, as well as good resistance to heavy metal exposure, with an order of metal tolerance determined as iron > copper > nickel > manganese > zinc > cadmium. Finally, we showed that the application of mineral fertilizer can in several cases enhance bacterial growth and thus potentially increase the relative proportion of the sporobiota in the olive grove ecosystem. In summary, the identification of the culturable olive sporobiota increases our understanding of the microbial diversity in Spanish olive groves, while tolerance and resistance profiles provide important insights into the phenotypic characteristics of the microbial community. IMPORTANCE Microbial communities are a key component of healthy soils. Spore-forming microorganisms represent a large fraction of this community-termed the "sporobiota"-and play a central role in creating a conducive environment for plant growth and food production. In addition, given their unique features, such as extraordinary stability and antimicrobial properties, members of the sporobiota present interesting candidates for biotechnological applications, such as sustainable plant protection products or in a clinical setting. For this, however, more information is needed on the spore-forming community of agricultural installations, ultimately promoting a transition toward a more sustainable agriculture.

Versatility of Stenotrophomonas maltophilia: Ecological roles of RND efflux pumps.

S. maltophilia is a widely distributed bacterium found in natural, anthropized and clinical environments. The genome of this opportunistic pathogen of environmental origin includes a large number of genes encoding RND efflux pumps independently of the clinical or environmental origin of the strains. These pumps have been historically associated with the uptake of antibiotics and clinically relevant molecules because they confer resistance to many antibiotics. However, considering the environmental origin of S. maltophilia, the ecological role of these pumps needs to be clarified. RND efflux systems are highly conserved within bacteria and encountered both in pathogenic and non-pathogenic species. Moreover, their evolutionary origin, conservation and multiple copies in bacterial genomes suggest a primordial role in cellular functions and environmental adaptation. This review is aimed at elucidating the ecological role of S. maltophilia RND efflux pumps in the environmental context and providing an exhaustive description of the environmental niches of S. maltophilia. By looking at the substrates and functions of the pumps, we propose different involvements and roles according to the adaptation of the bacterium to various niches. We highlight that i°) regulatory mechanisms and inducer molecules help to understand the conditions leading to their expression, and ii°) association and functional redundancy of RND pumps and other efflux systems demonstrate their complex role within S. maltophilia cells. These observations emphasize that RND efflux pumps play a role in the versatility of S. maltophilia.

Late Embryogenesis Abundant Proteins Contribute to the Resistance of Toxoplasma gondii Oocysts against Environmental Stresses.

Toxoplasma gondii oocysts, which are shed in large quantities in the feces from infected felines, are very stable in the environment, resistant to most inactivation procedures, and highly infectious. The oocyst wall provides an important physical barrier for sporozoites contained inside oocysts, protecting them from many chemical and physical stressors, including most inactivation procedures. Furthermore, sporozoites can withstand large temperature changes, even freeze-thawing, as well as desiccation, high salinity, and other environmental insults; however, the genetic basis for this environmental resistance is unknown. Here, we show that a cluster of four genes encoding Late Embryogenesis Abundant (LEA)-related proteins are required to provide Toxoplasma sporozoites resistance to environmental stresses. Toxoplasma LEA-like genes (TgLEAs) exhibit the characteristic features of intrinsically disordered proteins, explaining some of their properties. Our in vitro biochemical experiments using recombinant TgLEA proteins show that they have cryoprotective effects on the oocyst-resident lactate dehydrogenase enzyme and that induced expression in E. coli of two of them leads to better survival after cold stress. Oocysts from a strain in which the four LEA genes were knocked out en bloc were significantly more susceptible to high salinity, freezing, and desiccation compared to wild-type oocysts. We discuss the evolutionary acquisition of LEA-like genes in Toxoplasma and other oocyst-producing apicomplexan parasites of the Sarcocystidae family and discuss how this has likely contributed to the ability of sporozoites within oocysts to survive outside the host for extended periods. Collectively, our data provide a first molecular detailed view on a mechanism that contributes to the remarkable resilience of oocysts against environmental stresses. IMPORTANCE Toxoplasma gondii oocysts are highly infectious and may survive in the environment for years. Their resistance against disinfectants and irradiation has been attributed to the oocyst and sporocyst walls by acting as physical and permeability barriers. However, the genetic basis for their resistance against stressors like changes in temperature, salinity, or humidity, is unknown. We show that a cluster of four genes encoding Toxoplasma Late Embryogenesis Abundant (TgLEA)-related proteins are important for this resistance to environmental stresses. TgLEAs have features of intrinsically disordered proteins, explaining some of their properties. Recombinant TgLEA proteins show cryoprotective effects on the parasite's lactate dehydrogenase, an abundant enzyme in oocysts, and expression in E. coli of two TgLEAs has a beneficial effect on growth after cold stress. Moreover, oocysts from a strain lacking all four TgLEA genes were more susceptible to high salinity, freezing, and desiccation compared to wild-type oocysts, highlighting the importance of the four TgLEAs for oocyst resilience.

GF14f gene is negatively associated with yield and grain chalkiness under rice ratooning.

Background: Ratoon rice cropping has been shown to provide new insights into overcoming the current challenges of rice production in southern China. However, the potential mechanisms impacting yield and grain quality under rice ratooning remain unclear. Methods: In this study, changes in yield performance and distinct improvements in grain chalkiness in ratoon rice were thoroughly investigated, using physiological, molecular and transcriptomic analysis. Results: Rice ratooning induced an extensive carbon reserve remobilization in combination with an impact on grain filling, starch biosynthesis, and ultimately, an optimization in starch composition and structure in the endosperm. Furthermore, these variations were shown to be associated with a protein-coding gene: GF14f (encoding GF14f isoform of 14-3-3 proteins) and such gene negatively impacts oxidative and environmental resistance in ratoon rice. Conclusion: Our findings suggested that this genetic regulation by GF14f gene was the main cause leading to changes in rice yield and grain chalkiness improvement of ratoon rice, irrespective of seasonal or environmental effects. A further significance was to see how yield performance and grain quality of ratoon rice were able to be achieved at higher levels via suppression of GF14f.

Resistance of 3D-Printed Components, Test Specimens and Products to Work under Environmental Conditions-Review.

The development of additive manufacturing methods known as "3D printing" started in the 1980s. In these methods, spatial models are created from a semi-finished product such as a powder, filament or liquid. The model is most often created in layers, which are created from the semi-finished product, which is most often subjected to thermal treatment or using light or ultraviolet rays. The technology of additive manufacturing has both advantages and disadvantages when compared to the traditionally used methods of processing thermoplastic materials, such as, for example, injection or extrusion. The most important advantages are low cost, flexibility and speed of manufacturing of elements with different spatial shapes. From the point of view of the user of the product, the most important disadvantages are the lower mechanical properties and lower resistance to environmental factors that occur during the use of the manufactured products. The purpose of this review is to present current information and a compilation of features in the field of research on the effects of the interactions of different types of environments on the mechanical properties of 3D-manufactured thermoplastic products. Changes in the structure and mechanical properties of the material under the influence of factors such as humidity, salt, temperature, UV rays, gasoline and the environment of the human body are presented. The presented article enables the effects of environmental conditions on common materials used in 3D printing technology to be collated in one place.

Environmental Resistance and Fatigue Behaviors of Epoxy/Nano-Boron Nitride Thermally Conductive Structural Film Adhesive Toughened by Polyphenoxy.

The thermally conductive structural film adhesive not only carries large loads but also exhibits excellent heat-transfer performance, which has huge application prospects. Herein, a novel epoxy (Ep) thermally conductive structural film adhesive was prepared using polyphenoxy (PHO) as the toughening agent and film former, boron nitride (BN) nanosheets as the thermally conductive filler, and polyester fabric as the carrier. When the amount of PHO in the epoxy matrix was 30 phr and the content of nano-BN was 30 wt.% (Ep/PHO30/nBN30), the adhesive resin system showed good film-forming properties, thermal stability, and thermal conductivity. The glass transition temperature of Ep/PHO30/nBN30 was 215 °C, and the thermal conductivity was 209.5% higher than that of the pure epoxy resin. The Ep/PHO30/nBN30 film adhesive possessed excellent adhesion and peeling properties, and the double-lap shear strength at room temperature reached 36.69 MPa, which was 21.3% higher than that of pure epoxy resin. The double-lap shear strength reached 15.41 MPa at 150 °C, demonstrating excellent high temperature resistance. In addition, the Ep/PHO30/nBN30 film adhesive exhibited excellent heat-aging resistance, humidity, and medium resistance, and the shear strength retention rate after exposure to the complicated environment reached more than 90%. The structural film adhesive prepared showed excellent fatigue resistance in the dynamic load fatigue test, the double-lap shear strength still reached 35.55 MPa after 1,000,000 fatigue cycles, and the strength retention rate was 96.9%, showing excellent durability and fatigue resistance.

Dissection of the Activity of Agricultural Fungicides against Clinical Aspergillus Isolates with and without Environmentally and Medically Induced Azole Resistance.

Azole resistance is an emerging problem in patients with aspergillosis. The role of fungicides for resistance development and occurrence is not fully elucidated. EUCAST reference MICs of 17 fungicides (11 azoles and 6 others), five azole fungicide metabolites and four medical triazoles were examined against two reference and 28 clinical isolates of A. fumigatus, A. flavus and A. terreus with (n = 12) and without (n = 16) resistance mutations. Eight/11 azole fungicides were active against wild-type A. fumigatus, A. flavus and A. terreus, including four (metconazole, prothioconazole-desthio, prochloraz and imazalil) with low MIC50 (≤2 mg/L) against all three species and epoxiconazole, propiconazole, tebuconazole and difenoconazole also against wild-type A. terreus. Mefentrifluconazole, azole metabolites and non-azole fungicides MICs were >16 mg/L against A. fumigatus although partial growth inhibition was found with mefentrifluconazole. Moreover, mefentrifluconazole and axozystrobin were active against wild-type A. terreus. Increased MICs (≥3 dilutions) were found for TR34/L98H, TR34(3)/L98H, TR46/Y121F/T289A and G432S compared to wild-type A. fumigatus for epoxiconazole, propiconazole, tebuconazole, difenoconazole, prochloraz, imazalil and metconazole (except G432S), and for prothioconazole-desthio against TR46/Y121F/T289A, specifically. Increased MICs were found in A. fumigatus harbouring G54R, M220K and M220R alterations for five, one and one azole fungicides, respectively, compared to MICs against wild-type A. fumigatus. Similarly, increased MICs wer found for A. terreus with G51A, M217I and Y491H alterations for five, six and two azole fungicides, respectively. Azole fungicides showed activity against wild-type A. fumigatus, A. terreus and A. flavus, but not against all mutant isolates, suggesting the environmental route of azole resistance may have a role for all three species.

Preliminary Evaluation of In Vitro Bacteriostatic and Bactericidal Effect of Salt on Leptospira spp.

Environmental resistance is an important factor for understanding the epidemiology of leptospirosis. Recently, new Leptospira hosts were identified, including also marine mammals. Moreover, halotolerant Leptospira strain, isolated from the environment and animals, highlighted the capability of this microorganism to persist in the seawater. The aim of this research was to investigate the bacteriostatic and bactericidal effect of salt on Leptospira strains belonging to 16 different serovars. The minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) values were verified through the microdilutions method starting from a 20% sodium chloride concentration. MIC values obtained were between 0.3125% and 10% of salt, while MBC values between 0.625% and >20%. Icterohaemorrhagiae (MIC: 0.3125%; MBC: 0.625%) resulted the most inhibited serovar, while the most resistant was Tarassovi (MIC: 10%; MBC: >20%). Interestingly, trends were reported for Pomona (MIC: 1.25%; MBC: >20%) and Bratislava (MIC: 0.625%; MBC: 20%), highlighting low MIC values but high MBC values. This is the first investigation aimed at the in vitro effect of salt on the growth of Leptospira spp. reference strains.

Climate change and conservation in a warm North American desert: effect in shrubby plants.

BACKGROUND: Deserts are biologically rich habitats with a vast array of animals and plants adapted to xeric conditions, and most deserts are among the planet's last remaining areas of total wilderness. Among North American deserts, the Chihuahuan Desert has the highest levels of diversity and endemism. To understand the effect of future climate change on plants distributed in this arid land and propose effective conservation planning, we focused on five endemic shrubby species that characterize the Chihuahuan Desert and used an integrative approach. METHODS: Ecological niche-based modeling, spatial genetics and ecological resistance analyses were carried out to identify the effect of global warming on the studied five shrubby species. Key areas that need to be preserved were identified taking into account the existing protected areas within the Chihuahuan Desert. RESULTS: The extent of future distribution will vary among these species, and on average expansion will occur in the western part of the Chihuahuan Desert. For most species low environmental resistance to gene flow was predicted, while higher future resistance was predicted for one species that would lead to increased population isolation. The highest haplotype diversity was identified in three hotspots. Based on future suitability of habitat and in the haplotype diversity we suggest preserving two hotspots of genetic diversity in the Sierra Madre Oriental, located in areas without protection. The third hotspot was detected in the well preserved Tehuacán-Cuicatlán Man and Biosphere Reserve. CONCLUSION: Global climate change will have an effect in arid adapted plants, favoring expansion in the western of the Chihuahuan Desert however negatively affecting others with high ecological resistance disrupting gene flow. Two hotspots of genetic diversity in the Sierra Madre Oriental should be protected.

Recovery of Previously Uncultured Bacterial Genera from Three Mediterranean Sponges.

Sponges often harbour a dense and diverse microbial community. Presently, a large discrepancy exists between the cultivable bacterial fraction from sponges and the community in its natural environment. Here, we aimed to acquire additional insights into cultivability of (previously uncultured) bacteria from three sponge species, namely Aplysina aerophoba, Corticium candelabrum and Petrosia ficiformis, by studying bacterial growth on five media in the form of 60 communities scraped from plates without antibiotics, as well as in the form of individual isolates that were grown on these media supplemented with antibiotics. We applied (double-)barcoded 16S ribosomal RNA (rRNA) gene amplicon sequencing for species identification. We show that previously uncultured bacteria can be cultivated using conventional plating and that application of antibiotics in the media can serve to capture a greater bacterial diversity. Moreover, we present criteria to address an important caveat of the plate scraping method whereby bacteria may be detected that did not actually grow. Fourteen out of 27 cultivated novel taxa (<95% identity of the 16S rRNA gene amplicon to reported species) belong to Actinobacteria, which indicates the presence of a large untapped reservoir of bioactive compounds. Three Flavobacteriaceae spp. were isolated that potentially constitute two new genera and one new species.

Cross-Linked Protein Nanofilter with Antibacterial Properties for Multifunctional Air Filtration.

Development of high-performance nanomaterials with not only strong ability to trap the pollutants but also good structure stability under varying environmental conditions is a critical need for air-filtration applications. However, it has been very challenging for a filtering material to simultaneously realize multifunctional air filtration and good environmental stability. Here, based on our previous studies on protein-based nanofilters, we report a cross-linked protein nanofabric to address this challenging issue. It is found that cross-linked protein nanofabrics can significantly improve the structure stability against different moisture levels and temperatures, while maintaining the multifunctional filtration performance. Moreover, it is demonstrated that the cross-linked protein nanomaterials also possess antibacterial properties, such as Shewanella oneidensis bacteria, further improving the environmental stability. The effects of cross-linking with different loadings of cross-linking agent on the structure stability and filtration performance are further investigated at different humidity levels and temperatures. This study provides a cost-effective solution for advanced "green" nanomaterials with excellent performance in both filtration functions and structure stability under varying environment.

Functional metagenomic analysis reveals rivers are a reservoir for diverse antibiotic resistance genes.

The environment harbours a significant diversity of uncultured bacteria and a potential source of novel and extant resistance genes which may recombine with clinically important bacteria disseminated into environmental reservoirs. There is evidence that pollution can select for resistance due to the aggregation of adaptive genes on mobile elements. The aim of this study was to establish the impact of waste water treatment plant (WWTP) effluent disposal to a river by using culture independent methods to study diversity of resistance genes downstream of the WWTP in comparison to upstream. Metagenomic libraries were constructed in Escherichia coli and screened for phenotypic resistance to amikacin, gentamicin, neomycin, ampicillin and ciprofloxacin. Resistance genes were identified by using transposon mutagenesis. A significant increase downstream of the WWTP was observed in the number of phenotypic resistant clones recovered in metagenomic libraries. Common ß-lactamases such as blaTEM were recovered as well as a diverse range of acetyltransferases and unusual transporter genes, with evidence for newly emerging resistance mechanisms. The similarities of the predicted proteins to known sequences suggested origins of genes from a very diverse range of bacteria. The study suggests that waste water disposal increases the reservoir of resistance mechanisms in the environment either by addition of resistance genes or by input of agents selective for resistant phenotypes.

Novel resistance functions uncovered using functional metagenomic investigations of resistance reservoirs.

Rates of infection with antibiotic-resistant bacteria have increased precipitously over the past several decades, with far-reaching healthcare and societal costs. Recent evidence has established a link between antibiotic resistance genes in human pathogens and those found in non-pathogenic, commensal, and environmental organisms, prompting deeper investigation of natural and human-associated reservoirs of antibiotic resistance. Functional metagenomic selections, in which shotgun-cloned DNA fragments are selected for their ability to confer survival to an indicator host, have been increasingly applied to the characterization of many antibiotic resistance reservoirs. These experiments have demonstrated that antibiotic resistance genes are highly diverse and widely distributed, many times bearing little to no similarity to known sequences. Through unbiased selections for survival to antibiotic exposure, functional metagenomics can improve annotations by reducing the discovery of false-positive resistance and by allowing for the identification of previously unrecognizable resistance genes. In this review, we summarize the novel resistance functions uncovered using functional metagenomic investigations of natural and human-impacted resistance reservoirs. Examples of novel antibiotic resistance genes include those highly divergent from known sequences, those for which sequence is entirely unable to predict resistance function, bifunctional resistance genes, and those with unconventional, atypical resistance mechanisms. Overcoming antibiotic resistance in the clinic will require a better understanding of existing resistance reservoirs and the dissemination networks that govern horizontal gene exchange, informing best practices to limit the spread of resistance-conferring genes to human pathogens.