Analysis of temporal fecal microbiota dynamics in weaner pigs with and without exposure to enterotoxigenic Escherichia coli1,2

Muwonge

Hutchings

et al. 2020

Sci Rep

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Group antimicrobial administration is used to control disease in livestock, but we have little insight into how this impacts antimicrobial resistance (AMR) gene dynamics. Here, a longitudinal study was carried out during a single production cycle on a commercial pig unit with high historic and current antimicrobial usage. Quantitative PCR, 16S rRNA gene metabarcoding and shotgun metagenomic sequencing were used to track faecal AMR gene abundance and diversity and microbiome alpha diversity. Shotgun metagenomic sequencing identified 144 AMR genes in total, with higher AMR gene diversity present in young pigs compared to dry sows. Irrespective of in-feed antibiotic treatment or changes in microbiome diversity, mean AMR gene copy number was consistently high, with some AMR genes present at copy numbers comparable to the bacterial 16S rRNA gene. In conclusion, AMR gene prevalence and abundance were not influenced by antibiotic use, either during the production cycle or following whole-herd medication. The high levels of certain genes indicate they are widely disseminated throughout the microbial population, potentially aiding stability. Despite the high and relatively stable levels of resistance genes against the main antimicrobials used, these compounds continue to control production limiting diseases on this unit.Antimicrobial agents are used regularly in many agricultural systems worldwide to improve the health, welfare and productivity of livestock 1,2 . This has led to concerns about the anthropogenic selection of antimicrobial-resistant bacteria in livestock systems 2-4 , particularly due to the potential transfer of antimicrobial resistance (AMR) genes from livestock to humans 5-7 and into the environment 8,9 . In the United Kingdom, 52% of all antimicrobials sold for livestock were used in the pig and poultry sectors, with tetracyclines being reported as the highest sold antibiotic class 10 . Given the substantial use of antimicrobials in pig production, the association between antimicrobial use and AMR has been an area of intensive study. Specifically, antimicrobial administration has been associated with increased AMR in sentinel bacteria such as Escherichia coli [11][12][13] , Staphylococcus aureus 14,15 , and Campylobacter 16 , Salmonella 17 and Enterococcus 18 species. Molecular studies have revealed a higher richness and diversity of AMR genes in pigs administered oxytetracycline in-feed 19 , with farm origin being associated with AMR gene abundances in faeces at slaughter using both quantitative PCR 20 and metagenomic 21 datasets. However, there are currently few longitudinal studies of AMR in livestock systems, specifically for pig production 22 , with information on antimicrobial use in these systems often derived from national figures, rather than from farm medicines records 12,21 .Here, we examine microbial population dynamics and both AMR gene abundance and diversity on a single pig farm during a six-month production cycle. Faecal samples were collected weekly from a batch of young pigs that were ...

Section: Materials and Methods Study Farmmentioning

confidence: 99%

“…The generated sequences were processed using cutadapt 24 and mothur 25 (URL: https://www.mothur.org/wiki/ MiSeq_SOP. Accessed January 2018) as described previously 23 . Here, unique sequences were binned into operational taxonomic units (OTUs) using a database-independent approach.…”

Section: Materials and Methods Study Farmmentioning

confidence: 99%

Resistance to change: AMR gene dynamics on a commercial pig farm with high antimicrobial usage

Muwonge

Hutchings

et al. 2020

Sci Rep

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“…Faecal samples were collected from aged mice before and at 4 and 6 wk after passive microbiota transfer and from young donor mice. DNA was extracted and prepared for 16S rRNA gene sequencing, targeting the V3 hypervariable region, as described previously (Pollock et al, 2018). One library pool was constructed using equimolar concentrations of DNA from each of the included samples (n=39), as calculated using a fluorometric assay (Qubit dsDNA HS Assay kit, Thermo Fisher Scientific, Paisley, UK).…”

Section: Methodsmentioning

confidence: 99%

Microbial Stimulation Reverses the Age-Related Decline in M Cells in Aged Mice

Donaldson

Vohra

et al. 2020

Preprint

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SUMMARYAgeing has a profound effect on the immune system, termed immunosenescence, resulting in increased incidence and severity of infections and decreased efficacy of vaccinations. We previously showed that immunosurveillance in the intestine, achieved primarily through antigen sampling M cells in the follicle associated epithelium (FAE) of Peyer’s patches, was compromised during ageing due to a decline in M cell functional maturation. The intestinal microbiota also changes significantly with age, but whether this affects M cell maturation was not known. We show that housing of aged mice on used bedding from young mice, or treatment with bacterial flagellin, were each sufficient to enhance the functional maturation of M cells in Peyer’s patches. An understanding of the mechanisms underlying the influence of the intestinal microbiota on M cells has the potential to lead to new methods to enhance the efficacy of oral vaccination in aged individuals.

“…All samples collected from the sow barn and piglet accommodation were prepared for 16S rRNA gene metabarcoding targeting the V3 hypervariable region, as described in previous work (51). Six library pools were compiled using equimolar concentrations of DNA from each sample.…”

Section: Methodsmentioning

confidence: 99%

“…The generated sequences were processed using cutadapt (Martin, 2011) and mothur (Schloss et al, 2009) (URL: https://www.mothur.org/wiki/MiSeq_SOP. Accessed January 2018) as described previously (51). Here, unique sequences were binned using a database-independent approach.…”

Section: Methodsmentioning

confidence: 99%

Resistance to change? The impact of group medication on AMR gene dynamics during commercial pig production

Muwonge

Hutchings

et al. 2019

Preprint

Self Cite

The anthropogenic selection of antimicrobial resistance (AMR) genes is under intense scrutiny, particularly in livestock production, where group antimicrobial administration is used to control disease. Whilst large epidemiological studies provide important data on the diversity and distribution of AMR genes, we have little insight into how group antimicrobial administration impacts AMR gene abundance and diversity within a system. Here, faecal microbiome and AMR gene dynamics were tracked for six months through a standard production cycle on a commercial pig unit. Our results demonstrate that specific AMR genes have reached an equilibrium across this farming system to the extent that the levels measured were maintained from suckling through to slaughter, despite increases in microbiome diversity in early development. These levels were not influenced by antibiotic use, either during the production cycle or following whole herd medication. Some AMR genes were found at levels higher than that of the bacterial 16S rRNA gene, indicating widespread distribution across the most common bacterial genera. The targeted AMR genes were also detected in nearby soil samples, several with decreasing abundance with increasing distance from the unit, demonstrating that the farm acts as a point source of AMR gene ‘pollution’. Metagenomic sequencing of a subset of samples identified 144 AMR genes, with higher gene diversity in the piglet samples compared to the sow samples. Critically, despite overwhelming and stable levels of resistance alleles against the main antimicrobials used on this farm, these compounds continue to control the bacterial pathogens responsible for production losses and compromised welfare.ImportanceGroup antibiotic treatment has been used for decades to control bacterial diseases that reduce the productivity and compromise the welfare of livestock. Recent increases in antibiotic resistant infections in humans has resulted in concerns that antibiotic use in livestock may contribute to the development of untreatable bacterial infections in humans. There is however little understanding as to how the genes that bacteria require to become resistant to antibiotics respond during and after group antibiotic treatment of livestock, particularly in systems where high levels of antibiotics have been used for a prolonged period of time. We show that in such a system, levels of specific antibiotic resistance genes are high irrespective of group antibiotic treatments, whilst dramatic reductions in antibiotic use also fail to reduce the levels of these genes. These findings have important implications for public policy relating to the use of antibiotic in livestock farming.