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Antimicrobial resistance (AMR) is a global threat to human and animal health and well-being. To understand AMR dynamics, it is important to monitor resistant bacteria and resistance genes in all relevant settings. However, while monitoring of AMR has been implemented in clinical and veterinary settings, comprehensive monitoring of AMR in the environment is almost completely lacking. Yet, the environmental dimension of AMR is critical for understanding the dissemination routes and selection of resistant microorganisms, as well as the human health risks related to environmental AMR. Here, we outline important knowledge gaps that impede implementation of environmental AMR monitoring. These include lack of knowledge of the 'normal' background levels of environmental AMR, definition of high-risk environments for transmission, and a poor understanding of the concentrations of antibiotics and other chemical agents that promote resistance selection. Furthermore, there is a lack of methods to detect resistance genes that are not already circulating among pathogens. We conclude that these knowledge gaps need to be addressed before routine monitoring for AMR in the environment can be implemented on a large scale. Yet, AMR monitoring data bridging different sectors is needed in order to fill these knowledge gaps, which means that some level of national, regional and global AMR surveillance in the environment must happen even without all scientific questions answered. With the possibilities opened up by rapidly advancing technologies, it is time to fill these knowledge gaps. Doing so will allow for specific actions against environmental AMR development and spread to pathogens and thereby safeguard the health and wellbeing of humans and animals., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

Metagenomics can be used to monitor the spread of antibiotic resistance genes (ARGs). ARGs found in databases such as ResFinder and CARD primarily originate from culturable and pathogenic bacteria, while ARGs from non-culturable and non-pathogenic bacteria remain understudied. Functional metagenomics is based on phenotypic gene selection and can identify ARGs from non-culturable bacteria with a potentially low identity shared with known ARGs. In 2016, the ResFinderFG v1.0 database was created to collect ARGs from functional metagenomics studies. Here, we present the second version of the database, ResFinderFG v2.0, which is available on the Center of Genomic Epidemiology web server (https://cge.food.dtu.dk/services/ResFinderFG/). It comprises 3913 ARGs identified by functional metagenomics from 50 carefully curated datasets. We assessed its potential to detect ARGs in comparison to other popular databases in gut, soil and water (marine + freshwater) Global Microbial Gene Catalogues (https://gmgc.embl.de). ResFinderFG v2.0 allowed for the detection of ARGs that were not detected using other databases. These included ARGs conferring resistance to beta-lactams, cycline, phenicol, glycopeptide/cycloserine and trimethoprim/sulfonamide. Thus, ResFinderFG v2.0 can be used to identify ARGs differing from those found in conventional databases and therefore improve the description of resistomes., (© The Author(s) 2023. Published by Oxford University Press on behalf of Nucleic Acids Research.)

The knowledge of bacterial species diversity within the female urinary microbiome (FUM) is essential for understanding the role of the FUM in urinary tract health and disease. This study aimed to characterize the bacterial species diversity of the FUM of asymptomatic reproductive-age European women by combining extended culturomics and long-read sequencing of the near-full-length 16S rRNA gene. A total of 297 bacterial species (median of 53 species/sample) were identified, yet only 22% of the species were detected by both culture and sequencing methods. Recently recognized Gardnerella , Lactobacillus , and Limosilactobacillus species and 5 new putative Corynebacterium species were identified by culturomics, while anaerobic species (e.g., 11 Peptoniphilus spp.) were mostly detected by amplicon sequencing. Notably, there was not a single species common to all samples, although members of the genus Lactobacillus were detected in all. Lactobacillus crispatus, Lactobacillus iners, and Lactobacillus mulieris were observed in high relative abundance in several samples, as well as other species (e.g., Streptococcus agalactiae, Fannyhessea vaginae, Gardnerella vaginalis, Gardnerella swidsinskii), while low-abundance members (e.g., Finegoldia magna) were often more prevalent. A moderate correlation (Mantel test; r  = 0.5) between community structure types captured by culturomics and amplicon sequencing was observed, highlighting the benefit of combining both methodologies. This study provided a detailed FUM structure at the species level, which is critical to unveil the potential relationship between specific microbiome members and urinary diseases/disorders. Moreover, the different capacity to characterize microbiome profiles of culturomic and amplicon sequencing is described, providing valuable insights for further urinary microbiome studies. IMPORTANCE The bacterial species diversity within the female urinary microbiome (FUM) has been insufficiently characterized. This study demonstrated that complementarity between optimized culture-dependent and -independent approaches is highly beneficial for comprehensive FUM species profiling by detecting higher FUM species diversity than previously reported, including identification of unreported species belonging to the genera Lactobacillus , Limosilactobacillus , and Latilactobacillus and putative novel Corynebacterium species. Although some species were present in high relative abundance, low-abundance members were more prevalent. FUM classification into community structure types demonstrated high interindividual differences in urinary microbiome composition among asymptomatic women. We also report moderate correlation between culture-dependent and -independent derived data-highlighting drawbacks of each methodological approach. Our findings suggest that FUM bacterial diversity reported from previous studies may be underestimated. Finally, our results contribute to the fundamental knowledge of the FUM required for further exploration of the urinary microbiome role in urinary tract diseases.

Background: To date, information on healthy female urinary microbiota is available mostly at genus level and at one time point. However, profound species-level characterization of healthy urinary microbiome and its stability over time are essential for further correct interpretation of its role in healthy urogenital tract. In this study, we investigated female urogenital microbiome (FUM) at two timepoints (within 2.5-year interval) in young asymptomatic European women. We used culturomics with accurate isolates' identification (MALDI-TOF MS and gene markers sequencing) to understand species stability within healthy FUM., Results: Extended culturomics of voided midstream urine sample pairs revealed a mean Shannon diversity index of 1.25 and mean of 19 species/sample (range 5-39 species; total of 115 species; 1830 isolates). High overall species variability between individuals was captured by beta diversity and a variety of community structure types, with the largest cluster characterized by Lactobacillus crispatus, often in combination with Gardnerella vaginalis or Gardnerella genomospecies 3. Significant FUM composition differences, related to Finegoldia magna and Streptococcus anginosus, according to smoking status were found. A high species variability within individuals (Shannon index SD > 0.5 in 7 out of 10 sample pairs) with a mean of 29% of shared species (range 9.1-41.7%) was observed. Moreover, 4 out of 10 sample pairs clustered in the same community structure type. The stable FUM sample pairs presented high abundance of Lactobacillus crispatus, Streptococcus agalactiae or Lactobacillus paragasseri and Bifidobacterium spp.. Moreover, Gardnerella vaginalis, Gardnerella genomospecies 3 or Gardnerella swidsinskii were often maintained within individuals in high abundance., Conclusions: Shift in species composition at two distant timepoints was frequently observed among urogenital microbiome of European asymptomatic women. This suggests possible interchange of particular species in healthy FUM and the existence of multiple health-associated FUM compositions in certain individuals. Additionally, we provided additional evidence on resilience of particular bacterial communities and identified certain species more prone to persist in urogenital tract. This study revealed important details on the FUM composition complexity relevant for studies aiming to understand microbiota role in the urogenital tract health and for identification of eubiotic and dysbiotic FUM.

In order to face a constantly changing environment, animals need to be able to update their knowledge of the world on the basis of new information. Often, this means to inhibit a previously acquired response and flexibly change their behaviour to produce a new response. Here, we measured such abilities in young domestic chicks, employing a Colour Reversal Learning Task. During the acquisition phase, 17 one-week-old male chicks had to learn to peck on one of two coloured boxes to obtain a food reward. After reaching criterion, chicks underwent a reversal phase in which the previously learned colour-reward contingency was swapped. As expected from the literature, chicks performed better in the acquisition phase with respect to the reversal phase. Results moreover highlighted the presence of a lateralized bias selectively during reversal: chicks performed better if the stimulus rewarded was located in the left hemispace (processed by the right hemisphere). Interestingly, the bias correlated with the individual difficulty, i.e., it was stronger in those chicks which needed more trials to complete the reversal session. The present study contributes evidence in support of behavioural flexibility in young chicks, along with a novel perspective on lateralized mechanisms that might underlie such ability.