Determination and quantification of microbial communities and antimicrobial resistance on food through host DNA-depleted metagenomics

Bloomfield, Samuel; Zomer, Aldert; O’Grady, Justin; Kay, Gemma L.; Wain, John; Janecko, Nicol; Palau, Raphaëlle; Mather, Alison E.

doi:10.1016/j.fm.2022.104162

Cited by 17 publications

(23 citation statements)

References 35 publications

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“…Predominant Gram-negative bacteria like E. coli, P. aeruginosa , and K. pneumonia harboring diverse resistance mechanisms have been previously identified as common sepsis-causing pathogens (Bloomfield et al, 2023; Mu et al, 2023). Some of these species have been linked to gut-to-lung bacterial migration and AMR acquisition and the worsening of sepsis (Wheatley et al, 2022).…”

Section: Discussionmentioning

confidence: 99%

Tracheal aspirate metagenomics reveals association of antibiotic resistance with non-pulmonary sepsis mortality

Rodríguez-Pérez,

Ciuffreda,

Hernández-Beeftink

et al. 2024

Preprint

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Background. Previous metabarcoding studies based on 16S rRNA sequencing in patients with extrapulmonary sepsis have found early pulmonary dysbiosis associated with a poor prognosis. To further discern this association, here we aimed to better characterize the pulmonary bacterial communities in these patients by leveraging metagenomics and to evaluate if the presence of antibiotic resistance genes (ARGs) could explain the higher mortality of the patients. Material and methods. Metagenomic sequencing was performed using the Nextera XT Library Prep Kit and HiSeq 4000 (Illumina Inc.) on tracheal aspirate samples that were obtained within 24 h from diagnosis from patients with extrapulmonary sepsis admitted to the Intensive Care Unit (ICU). Analysis involved MetaSpades for contig assembly, Kraken2 and Metaphlan4 for taxonomic classification, and CARD and GTDB-tk for ARGs annotation and assignment to the bacterial species. The relationship between the presence of antibiotic resistance and ICU mortality was evaluated using the Wilcoxon test and logistic regression models adjusting for clinical and demographic variables. Results. In total, 127 different ARGs were detected circumscribed only to seven patients. The most common ARGs found were from the antibiotic groups of aminoglycosides and beta-lactams, both present in most of the patients. These ARGs were found, almost entirely linked to Klebsiella pneumoniae, Escherichia coli, and Pseudomonas aeruginosa. The results also show a significant enrichment of ARGs among patients who died while admitted in the ICU (57%, 95% confidence interval [CI]: 18-90%) compared to surviving patients (20%, 95% CI: 7-40%) (p=0.022). Analyses adjusting for clinical and demographic variables did not alter this result. Conclusion. Metagenomic sequencing has allowed an unprecedented characterization of the sepsis lung microbiome showing that antibiotic resistance is common among these patients. The results also suggest a relationship between the early accumulation of ARGs in the lung of patients with extrapulmonary sepsis who die while admitted in the ICU. Studies in independent samples will be needed to validate our findings.

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Section: Discussionmentioning

confidence: 99%

Tracheal aspirate metagenomics reveals association of antibiotic resistance with non-pulmonary sepsis mortality

Rodríguez-Pérez,

Ciuffreda,

Hernández-Beeftink

et al. 2024

Preprint

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“…and Psychrobacter spp. are frequently the dominant taxa in a variety of food processing environments [55,58,59,64] due to their tolerance to low temperatures [68] and their ability to form biofilms [69,70] which often results in contamination of food products [71,72]. To our knowledge, the presence of Sphingomonas spp.…”

Section: Discussionmentioning

confidence: 99%

Microbial composition and dynamics in environmental samples from a ready-to-eat food production facility with a long-term colonisation of Listeria monocytogenes

Maria,

Aird,

Viet

et al. 2024

Preprint

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Background: Listeria monocytogenes is a foodborne pathogen of significant concern for the food industry due to the potential health threat to consumers and a remarkable ability to persist through food safety control efforts. Even with rigorous cleaning and disinfection practices in food production settings, L. monocytogenes and resident microbes can thrive and then potentially spillover from the environment to foods. Understanding the composition and interactions within microbial communities in food processing environments can reveal mechanisms on L. monocytogenes survival and persistence in these settings. In this study, we used metagenomic analysis to monitor the resident microbiota in non-food contact surfaces located in the preparation and production areas of the high care zone of a facility producing ready-to-eat foods that was previously known to have a persistent ST121 strain of L. monocytogenes. Results: A consistent composition and relative abundance of most microbial taxa over sampling sites and times were observed in environmental metagenomes, with the genus Pseudomonas (principally Pseudomonas fluorescens) representing a major constituent of the metagenome. While there were highly related populations between the production and preparation areas, individual taxa characteristic of each area were observed (e.g., Sphingomonas aerolata was more abundant in the production area). Notably Listeria spp. were not observed in the metagenomes, but were detected in cultured samples, suggesting the abundance of Listeria was below the detection threshold for direct metagenomics. As such, a quasimetagenomics approach (sequencing of microbiological culture enrichments) was used to detect and subtype Listeria spp. and L. monocytogenes. While possible to classify Listeria seeligeri sequence types using quasimetagenomes, the classification of the L. monocytogenes sequence types was limited to a low proportion of the samples. Conclusions: A stable resident microbiota was observed with multiple environmental-associated microbial taxa, emphasizing that non-contact food surfaces represent a niche that can be colonised by a community of interdependent microbes. This community was likely selected for by their collective ability to survive cleaning and disinfection control efforts in this environment. Listeria spp. was a member of this microbial community, albeit at very low abundance, and likewise may be benefiting from the mutualism of the overall microbial community.

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“…Our approach is unique among the latter, which have a longer turnaround time and associated costs (Charre et al 2020;Quéromès et al 2021;Bal et al 2020). In addition, other studies have used nanopore AS in different contexts, with SARS-CoV-2, e.g., combined with PCR barcoding protocol from ONT (with 25 PCR cycles) to improve the recovery of SARS-CoV-2 viral genomes in ten clinical samples (Lin et al 2022) or with customized sets of long amplicons (M. Wang et al 2020), but also to enrich for specific bacterial species (Marquet et al 2022;Martin et al 2022;Gan et al 2021;Payne et al 2021;Bloomfield et al 2023), but none have applied AS with successful SARS-CoV-2 whole genome sequencing without prior amplification.…”

Section: Discussionmentioning

confidence: 99%

NASCarD (Nanopore Adaptive Sampling with Carrier DNA): A rapid, PCR-free method for whole genome sequencing of pathogens in clinical samples

Miani

Borcard

Gempeler

et al. 2023

Preprint

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Whole-genome sequencing (WGS) represents the main technology for SARS-CoV-2 lineage characterization in diagnostic laboratories worldwide. The rapid, near-full-length sequencing of the viral genome is commonly enabled by high-throughput sequencing of PCR amplicons derived from cDNA molecules. Here, we present a new approach, called NASCarD (Nanopore adaptive sampling with carrier DNA), which allows low amount of nucleic acids to be sequenced while selectively enriching for sequences of interest, hence limiting the production of non-target sequences. Using clinical samples positive for SARS-CoV-2 during the Omicron wave, we demonstrate how the method leads to up to >100x coverage of the full genome sequences of the target organism as compared to standard shotgun metatranscriptomics approach. It provides complete and accurate genome sequence reconstruction within seven hours at a competitive cost. The new approach may have applications beyond SARS-CoV-2 sequencing for other DNA or RNA pathogens in clinical samples.

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Determination and quantification of microbial communities and antimicrobial resistance on food through host DNA-depleted metagenomics

Cited by 17 publications

References 35 publications

Tracheal aspirate metagenomics reveals association of antibiotic resistance with non-pulmonary sepsis mortality

Tracheal aspirate metagenomics reveals association of antibiotic resistance with non-pulmonary sepsis mortality

Microbial composition and dynamics in environmental samples from a ready-to-eat food production facility with a long-term colonisation of Listeria monocytogenes

NASCarD (Nanopore Adaptive Sampling with Carrier DNA): A rapid, PCR-free method for whole genome sequencing of pathogens in clinical samples

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