Academic Journal
Optimizing a culture-enriched hybrid metagenomics pipeline to assess the AMR footprint of livestock manure in anaerobic digestate.
| Τίτλος: | Optimizing a culture-enriched hybrid metagenomics pipeline to assess the AMR footprint of livestock manure in anaerobic digestate. |
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| Συγγραφείς: | Rahman N; Department of Animal Science, University of Manitoba, Winnipeg, Manitoba, Canada., Rahman ASMZ; Department of Animal Science, University of Manitoba, Winnipeg, Manitoba, Canada., Levin DB; Department of Biosystems Engineering, University of Manitoba, Winnipeg, Manitoba, Canada., McAllister TA; Department of Animal Science, University of Manitoba, Winnipeg, Manitoba, Canada.; Lethbridge Research Centre, Agriculture and Agri-Food Canada, Lethbridge, Alberta, Canada., Cicek N; Department of Biosystems Engineering, University of Manitoba, Winnipeg, Manitoba, Canada., Derakhshani H; Department of Animal Science, University of Manitoba, Winnipeg, Manitoba, Canada. |
| Πηγή: | Microbiology spectrum [Microbiol Spectr] 2026 Oct 06; Vol. 14 (10), pp. e0247226. Date of Electronic Publication: 2026 Sep 03. |
| Τύπος έκδοσης: | Journal Article |
| Γλώσσα: | English |
| Στοιχεία περιοδικού: | Publisher: ASM Press Country of Publication: United States NLM ID: 101634614 Publication Model: Print-Electronic Cited Medium: Internet ISSN: 2165-0497 (Electronic) Linking ISSN: 21650497 NLM ISO Abbreviation: Microbiol Spectr Subsets: MEDLINE |
| Imprint Name(s): | Original Publication: Washington, DC : ASM Press, 2013- |
| Ιατρικοί όροι (MeSH): | Manure*/microbiology , Metagenomics*/methods , Livestock*/microbiology , Bacteria*/genetics , Bacteria*/drug effects , Bacteria*/isolation & purification , Bacteria*/classification , Drug Resistance, Bacterial*/genetics, Anti-Bacterial Agents/pharmacology ; Animals ; Anaerobiosis ; Interspersed Repetitive Sequences |
| Περίληψη: | The role of environmental samples from livestock production systems, including manure and anaerobic digestate, as reservoirs of antimicrobial resistance genes (ARGs) is likely underestimated because conventional metagenomic approaches can overlook low-abundance ARGs and often lack the resolution to associate these genes with their microbial hosts and co-localized mobile genetic elements (MGEs). We evaluated whether culture-enriched metagenomics (CEMG), with and without antibiotic selection, enhances ARG detection in anaerobic digestate and improves the resolution of ARG-MGE-host associations using hybrid short- and long-read metagenomic assembly. CEMG increased ARG recovery; mean ARG abundance rose from 15.4 counts per million (CPM) in metagenomic fresh digestate (FD) to 124 CPM in CEMG without antibiotics and 160 CPM in antibiotic-selective CEMG. In FD, only 9 unique ARGs were detected, whereas CEMG recovered 112, including ARGs of clinical importance, such as glycopeptide resistance, beta-lactamase genes, and the cfr 23S rRNA methyltransferase conferring cross-resistance to multiple antibiotic classes. Antibiotic selection induced targeted, class-specific shifts in ARG profiles, with ARGs associated with tetracycline resistance consistently enriched across treatments. Hybrid metagenomic assembly resolved the genomic context of 784 ARGs, of which 59.3% were co-localized with at least one class of MGEs, predominantly plasmids and integrative conjugative elements/integrative mobilizable elements. Biocide and metal resistance genes frequently co-occurred with ARGs on the same contigs. Together, these findings demonstrate that antibiotic-selective culture enrichment enhances resistome surveillance by improving detection of low-abundance ARGs, while hybrid assembly provides critical genomic context for assessing their mobility and host associations.IMPORTANCELivestock manure and its byproducts, such as anaerobic digestate, are recognized as important environmental reservoirs of antimicrobial resistance genes (ARGs) and resistant bacteria, yet current metagenomic approaches may underestimate this risk by failing to detect low-abundance but clinically relevant ARGs. Here, we show that integrating culture enrichment with hybrid metagenomics improves ARG recovery and reveals ARG co-localization with mobile genetic elements and putative bacterial hosts. This approach captures a cultivable and condition-responsive fraction of the resistome that is not readily accessible through direct metagenomic sequencing alone, providing a more informative framework for environmental AMR surveillance. |
| Competing Interests: | The authors declare no conflict of interest. |
| Grant Information: | RGPIN-2023-04359 Natural Sciences and Engineering Research Council of Canada |
| Contributed Indexing: | Keywords: anaerobic digestion; antimicrobial resistance; culture-enriched metagenomics; livestock manure |
| Substance Nomenclature: | 0 (Manure) 0 (Anti-Bacterial Agents) |
| Entry Date(s): | Date Created: 20260903 Date Completed: 20261007 Latest Revision: 20261007 |
| Update Code: | 20261007 |
| DOI: | 10.1128/spectrum.02472-26 |
| PMID: | 42690060 |
| Βάση Δεδομένων: | MEDLINE |
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