Antibiotics' action, once a ‘magic bullet’, is now hindered by widespread microbial resistance, creating a global antimicrobial resistance (AMR) crisis. A primary driver of AMR is the selective pre... Plain language summary Antibiotics are becoming less effective at fighting infections because of antimicrobial resistance (AMR). This phenomenon is mainly caused by the abuse and misuse of antibiot...

Recycled manure solids (RMS) are increasingly adopted in dairy farming for their economic advantages and their role in improving nutrient recycling and waste management; however, concerns regarding greenhouse gas (GHG) emissions during storage persist. This study assessed the effects of biochar supplementation at 2.5% (2.5B) and 10% (10B) compared to untreated RMS (C−) and acidified RMS (C+) on GHG emissions (measured both continuously and intermittently) and RMS characteristics during a one-month storage period. The results showed that the addition of biochar increased heavy metals concentration (with the exception of molybdenum) and the electrical conductivity of the RMS. Storage of RMS generally led to an increase in its dry matter content, except in the 10B treatment. The results showed that 10% biochar significantly reduced cumulative CO₂ and N₂O emissions, resulting in a 32% GWP reduction compared to untreated RMS. In contrast, the 2.5% dose led to higher CO₂ emissions, possibly due to microbial stimulation. Adding 10% biochar mitigated GHG emissions similarly to H₂SO₄ acidification but with fewer environmental and operational risks, making it a preferable farm-scale option. Continuous monitoring captured transient emission peaks, highlighting the importance of high-resolution assessments. Despite the emissions generated during biochar production, its application in RMS bedding systems offsets these environmental costs by mitigating GHG emissions and increasing nutrient content. Biochar’s mitigation potential, especially at higher doses, presents a safer, multifunctional alternative that aligns with EU climate goals. These findings support the integration of biochar into sustainable manure management strategies, though further research is needed to optimize application rates and assess cost-effectiveness in dairy farming. However, continued assessments at a larger scale and with different biochar addition rates are necessary to fully determine the potential of biochar supplementation to RMS.

The widespread use of Recycled Manure Solids (RMS) as cow bedding material is not without risks, since cattle manure may act as a vehicle for pathogenic and antimicrobial resistant bacteria dissemination. Thus, our aim was to evaluate RMS-supplemented with a pine biochar produced in Portugal as a new cow bedding material, since the use of biochar has been shown to have the potential to mitigate the impact of relevant bacterial species when added to animal manure microbiota. Our experimental setup consisted on fresh RMS samples that were collected on a commercial dairy farm and placed in naturally-ventilated containers for a total of 4 groups: 1-non-supplemented RMS; 2-RMS supplemented with 2.5% (wt/wt) of biochar; 3-RMS supplemented with 5% (wt/wt) of biochar; and 4-RMS supplemented with 10% (wt/wt) of biochar. Sampling was performed at 4 different incubation times (0, 5, 15 and 30 d) and in 2 distinct seasons: April-May (humid season) and June-July (dry season). The resulting 32 samples were subjected to DNA extraction and their microbiome profile determined through complete 16S rDNA gene sequencing using Nanopore next-generation sequencing. We observed that biochar supplementation clearly altered the microbiome of RMS, which was reflected in changes in populations' diversity and their relative abundance of relevant pathogenic bacteria. In particular, we found that long-term storage (30 d) was more beneficial than short-term storage, an effect that was more evident for samples supplemented with 2.5% or 5% biochar. In both seasons, those concentrations of biochar led to a decrease in the levels of several mastitis-causing agents (Enterobacteriaceae, streptococci, enterococci and staphylococci). In addition, we also observed a reduction in the levels of Salmonella spp. and Gram-positive bacilli in the biochar-supplemented samples. Unexpectedly, however, those same conditions yielded an increase in the abundance of Brucella spp., a group which includes important infectious agents, highlighting the need for a deeper evaluation of the impact of biochar supplementation of RMS to ensure the future safe and sustainable use of this environmental-friendly resource in animal production.

The use of recycled manure solids (RMS) as cow bedding in dairy farms poses concerns due to its potential to harbor pathogenic and antimicrobial-resistant bacteria. This study evaluated the impact of RMS supplementation with biochar at three concentrations (2.5%, 5%, and 10%) on bacterial counts and on the antimicrobial resistance and virulence profiles of Escherichia coli and Enterococcus isolates. The results show that biochar supplementation did not promote a significant reduction in bacterial numbers. Overall, there were no significant changes in the isolates’ virulence or resistance profiles, and none of the isolates obtained were classified as high or moderate threats based on their MAR and VIR indexes. The most pathogenic Enterococcus isolates found were present in the control samples. A phylogenetic analysis of E. coli isolates allowed us to identify phylogroup D strains, predominantly in RMS supplemented with 2.5% and 10% biochar, which are associated with higher virulence and resistance. These findings indicate that the biochar concentrations tested were not effective in significantly reducing the bacterial risks associated with RMS. Further research is needed to evaluate different biochar formulations and concentrations, aiming to optimize its potential use for RMS supplementation.

Ruminant herbivores meet their carbon and energy requirements from a symbiotic relationship with cellulosome-producing anaerobic bacteria that efficiently degrade plant cell-wall polysaccharides. The assembly of carbohydrate-active enzymes (CAZymes) into cellulosomes enhances protein stability and enzyme synergistic interactions. Cellulosomes comprise diverse CAZymes displaying a modular architecture in which a catalytic domain is connected,vialinker sequences, to one or more noncatalytic carbohydrate-binding modules (CBMs). CBMs direct the appended catalytic modules to their target substrates, thus facilitating catalysis. The genome of the ruminal cellulolytic bacteriumRuminococcus flavefaciensstrain FD-1 contains over 200 modular proteins containing the cellulosomal signature dockerin module. One of these is an endoglucanase Cel5A comprising two family 5 glycoside hydrolase catalytic modules (GH5) flanking an unclassified CBM (termed CBM-Rf2) and a C-terminal dockerin. This novel CBM-Rf2 has been purified and crystallized, and data from cacodylate-derivative crystals were processed to 1.02 and 1.29 Å resolution. The crystals belonged to the orthorhombic space groupP2₁2₁2₁. The CBM-Rf2 structure was solved by a single-wavelength anomalous dispersion experiment at the As edge.