Metagenomic Comparison of Antibiotic Resistance Genes Associated with Liquid and Dewatered Biosolids

Abstract

Municipal biosolids (MBs) that are land-applied in North America are known to possess an active microbial population that can include human pathogens. Activated sludge is a hotspot for the accumulation of antibiotics and has been shown to be a selective environment for microorganisms that contain antibiotic resistance genes (ARGs); however, the prevalence of ARGs in MBs is not well characterized. In this study, we enriched the plasmid metagenome from raw sewage sludge and two CP2 MBs, a mesophilic anaerobic digestate and a dewatered digestate, to evaluate the presence of ARGs in mobile genetic elements. The CP2-class biosolids are similar to Class B biosolids in the United States. The CP2 biosolids must meet a microbiological cut off of 2 × 10 colony-forming units (CFU) per dry gram or 100 mL of biosolids. The enriched plasmid DNA was sequenced (Illumina MiSeq). Sequence matching against databases, including the Comprehensive Antibiotic Resistance Database (CARD), MG-RAST, and INTEGRALL, identified potential genes of interest related to ARGs and their ability to transfer. The presence and abundance of different ARGs varied between treatments with heterogeneity observed among the same sample types. The MBs plasmid-enriched metagenomes contained ARGs associated with resistance to a variety of antibiotics, including β-lactams, rifampicin, quinolone, and tetracycline as well as the detection of extended spectrum β-lactamase genes. Cultured bacteria from CP2 MBs possessed antibiotic resistances consistent with the MBs metagenome data including multiantibiotic-resistant isolates. The results from this study provide a better understanding of the ARG and MGE profile of the plasmid-enriched metagenome of CP2 MBs.

Early-life gut microbiome modulation reduces the abundance of antibiotic-resistant bacteria

“…The final taxonomic affiliation analysis of the ARGs was performed with a MEtaGenome Analyzer (MEGAN v6.9) [44], which was used to compute and interactively explore the taxonomic content of the data set using the lowest common ancestor (LCA) method on the NCBI Taxonomy phylogenetic Tree of Life (LCA parameters: mini-score 35, top percentage 10%). The portion of sequence types or subtypes identified as ARGs in the total metagenome were defined as the relative abundance, using cross-sample normalization in ppm (one read in one million mapped reads) [42, 45].…”

Antimicrobal Resistance: Fecal Sanitation Strategies for Combatting a Global Public Health Threat

“…In this manner, a profile of the types and numbers of ARGs detected in a sample can be obtained and compared with other samples using various bioinformatics techniques and graphical representations. Metagenomics has been successfully applied in this manner for monitoring ARGs in wastewater treatment plants (Schluter et al, 2008;Wang et al, 2013;Yang et al, 2014;Li et al, 2015a;Munck et al, 2015;Zhang et al, 2015a;Bengtsson-Palme et al, 2016;Hu et al, 2016;Karkman et al, 2016;Ma et al, 2016b), biosolids (McCall et al, 2016;Tao et al, 2016;Rowe et al, 2016;Tang et al, 2016), manure (Agga et al, 2015), soil (Yan et al, 2016), rivers (Garner et al, 2016;Rowe et al, 2016), sediments (Cummings et al, 2011), and (Port et al, 2012). However, next-generation DNA sequencing technologies are still costly and require a high level of expertise, currently restricting metagenomic analysis to the realm of research, although that may soon change.…”

Genomic and functional techniques to mine the microbiome for novel antimicrobials and antimicrobial resistance genes

“…However, it is not suited for the detection of point mutations in chromosomal target genes known to be associated with antimicrobial resistance genes . Many studies have annotated antimicrobial resistance using the CARD alone or in combination with the ARDB . Unlike the ARDB, the CARD is actively managed and periodically updated.…”

“…120 Many studies have annotated antimicrobial resistance using the CARD alone or in combination with the ARDB. 110,111,121,122 Unlike the ARDB, the CARD is actively managed and periodically updated.…”