Biology subjects

Barker, L.

Publications and source records attributed to Barker, L..

8 recordsLinked to original sources

bioRxiv2026

Monkey hear, monkey do what? An application of Automated Behavioural Response systems for hypothesis testing in the worlds smallest monkey

Observer effects are a frequent problem in animal behaviour studies, particularly when assessing responses to human disturbance. Automated Behavioural Response (ABR) systems, which combine camera traps with automated sound playbacks, offer a solution but have been primarily used on large terrestrial mammals. Here, we demonstrate their use in a small ([~]110g) arboreal primate, the eastern pygmy marmoset (Cebuella niveiventris). We conducted two playback experiments to test the risk-disturbance and distracted prey hypotheses. The marmosets exhibited strong anti-predator responses to avian predator calls, including increased fleeing and vocalisations. Human speech elicited similar but weaker responses, indicating that pygmy marmosets do not perceive raptors and humans as equivalent threats. Embedding predator calls into anthropogenic noise reduced vocal responses, suggesting that anthropogenic noise interferes with responses to predation cues. Across five weeks, we generated 128 successful experimental trials, demonstrating that ABRs can rapidly produce sample sizes sufficient for hypothesis testing in the field.

animal behavior and cognition↗

bioRxiv2022

Plasmid overlap and evolution between Enterobacterales isolates from bloodstream infections and non-human compartments

Plasmids enable the dissemination of antimicrobial resistance (AMR) in common Enterobacterales pathogens, representing a major public health challenge. However, the extent of plasmid sharing and evolution between Enterobacterales causing human infections and other niches remains unclear, including the emergence of resistance plasmids. Dense, unselected sampling is highly relevant to developing our understanding of plasmid epidemiology and designing appropriate interventions to limit the emergence and dissemination of plasmid-associated AMR. We established a geographically and temporally restricted collection of human bloodstream infection (BSI)-associated, livestock-associated (cattle, pig, poultry, and sheep faeces, farm soils) and wastewater treatment work (WwTW)-associated (influent, effluent, waterways upstream/downstream of effluent outlets) Enterobacterales. Isolates were collected between 2008-2020 from sites <60km apart in Oxfordshire, UK. Pangenome analysis of plasmid clusters revealed shared "backbones", with phylogenies suggesting an intertwined ecology where well-conserved plasmid backbones carry diverse accessory functions, including AMR genes. Many plasmid "backbones" were seen across species and niches, raising the possibility that plasmid movement between these followed by rapid accessory gene change could be relatively common. Overall, the signature of identical plasmid sharing is likely to be a highly transient one, implying that plasmid movement might be occurring at greater rates than previously estimated, raising a challenge for future genomic One Health studies. FundingThis study was funded by the Antimicrobial Resistance Cross-council Initiative supported by the seven research councils and the NIHR, UK.

genomics↗

bioRxiv2020

Genomic network analysis of an environmental and livestock IncF plasmid population

IncF plasmids are diverse and of great clinical significance, often carrying genes conferring antimicrobial resistance (AMR) such as extended-spectrum {beta}-lactamases, particularly in Enterobacteriaceae. Organising this plasmid diversity is challenging, and current knowledge is largely based on plasmids from clinical settings. Here, we present a network community analysis of a large survey of IncF plasmids from environmental (influent, effluent, and upstream/downstream waterways surrounding wastewater treatment works) and livestock settings. We use a tractable and scalable methodology to examine the relationship between plasmid metadata and network communities. This reveals how niche (sampling compartment and host genera) partition and shape plasmid diversity. We also perform pangenome-style analyses on network communities. We show that such communities define unique combinations of core genes, with limited overlap. Building plasmid phylogenies based on alignments of these core genes, we demonstrate that plasmid accessory function is closely linked to core gene content. Taken together, our results suggest that stable IncF plasmid backbone structures can persist in environmental settings while allowing dramatic variation in accessory gene content that may be linked to niche adaptation. The recent association of IncF plasmids with AMR likely reflects their suitability for rapid niche adaptation.

bioinformatics↗

bioRxiv2020

Niche and local geography shape the pangenome of wastewater- and livestock-associated Enterobacteriaceae

Escherichia coli and other Enterobacteriaceae are highly diverse species with open pangenomes1,2, where genes move intra- and inter-species via horizontal gene transfer3. These species can cause clinical infections4,5 as well as persist environmentally6,7. Environmental populations have been suggested as important reservoirs of antimicrobial resistance (AMR) genes. However, as most analyses focus on clinical isolates8,9, the pangenome dynamics of natural populations remain understudied, particularly the role of plasmids. Here, we reconstructed near-complete genomes for 828 Enterobacteriaceae, including 553 Escherichia spp. and 275 non-Escherichia species with 2,293 circularised plasmids in total, collected from nineteen locations (livestock farms and wastewater treatment works in the United Kingdom) within a 30km radius at three timepoints over the course of a year. We find different dynamics for the chromosomal and plasmid-borne components of the pangenome, showing that plasmids have a higher burden of both AMR genes and insertion sequences, and AMR plasmids show evidence of being under stronger selective pressure. Focusing on E. coli, we observe that plasmid dynamics are more strongly dominated by niche and local geography, rather than phylogeny or season. Our results highlight the diversity of the AMR reservoir in these species and niches, and the importance of local strategies for controlling the emergence and spread of AMR.

microbiology↗

bioRxiv2020

Optimised use of Oxford Nanopore Flowcells for Hybrid Assemblies

Hybrid assemblies are highly valuable for studies of Enterobacteriaceae due to their ability to fully resolve the structure of mobile genetic elements, such as plasmids, which are involved in the carriage of clinically important genes (e.g. those involved in AMR/virulence). The widespread application of this technique is currently primarily limited by cost. Recent data has suggested that non-inferior, and even superior, hybrid assemblies can be produced using a fraction of the total output from a multiplexed nanopore (Oxford Nanopore Technologies [ONT]) flowcell run. In this study we sought to determine the optimal minimal running time for flowcells when acquiring reads for hybrid assembly. We then evaluated whether the ONT wash kit might allow users to exploit shorter running times by sequencing multiple libraries per flowcell. After 24 hours of sequencing, most chromosomes and plasmids had circularised and there was no benefit associated with longer running times. Quality was similar at 12 hours suggesting shorter running times are likely to be acceptable for certain applications (e.g. plasmid genomics). The ONT wash kit was highly effective in removing DNA between libraries. Contamination between libraries did not appear to affect subsequent hybrid assemblies, even when the same barcodes were used successively on a single flowcell. Utilising shorter run-times in combination with between-library nuclease washes allows at least 36 Enterobacteriaceae isolates to be sequenced per flowcell, significantly reducing the per isolate sequencing cost. Ultimately this will facilitate large-scale studies utilising hybrid assembly advancing our understanding of the genomics of key human pathogens. Data SummaryO_LIRaw sequencing data is available via NCBI under project accession number PRJNA604975. Sample accession numbers are provided in table S1. C_LIO_LIAssemblies are available via Figshare https://doi.org/10.6084/m9.figshare.11816532.v1. C_LI Impact StatementMost existing sequencing data has been acquired from short-read platforms (eg. Illumina). For some species of bacteria, clinically important genes, such as those involved in antibiotic resistance and/or virulence, are carried on plasmids. Whilst Illumina sequencing is highly accurate, it is generally unable to resolve complete genomic structures due to repetitive regions. Hybrid assembly uses long reads to scaffold together short-read contigs, maximising the benefits of both technologies. A major limiting factor to using hybrid assemblies at scale is the cost of sequencing the same isolate with two different technologies. Here we show that high-quality hybrid assemblies can be created for most isolates using significantly shorter run-times than are currently standard. We demonstrate that a simple washing step allows several libraries to be run on the same flowcell, facilitating the ability to take advantage of shorter running times. Adding nuclease means that contamination between libraries is minimal and has no significant effect on the quality of subsequent hybrid assemblies. This approach reduces the cost of acquiring long reads by >30%, paving the way for large-scale studies utilising hybrid assemblies which will likely significantly enhance our understanding of the genomics of important human pathogens.

bioinformatics↗

bioRxiv2020

High precision Neisseria gonorrhoeae variant and antimicrobial resistance calling from metagenomic Nanopore sequencing

The rise of antimicrobial resistant Neisseria gonorrhoeae is a significant public health concern. Against this background, rapid culture-independent diagnostics may allow targeted treatment and prevent onward transmission. We have previously shown metagenomic sequencing of urine samples from men with urethral gonorrhoea can recover near-complete N. gonorrhoeae genomes. However, disentangling the N. gonorrhoeae genome from metagenomic samples and robustly identifying antimicrobial resistance determinants from error-prone Nanopore sequencing is a substantial bioinformatics challenge. Here we demonstrate an N. gonorrhoeae diagnostic workflow for analysis of metagenomic sequencing data obtained from clinical samples using R9.4.1 Nanopore sequencing. We compared results from simulated and clinical infections with data from known reference strains and Illumina sequencing of isolates cultured from the same patients. We evaluated three Nanopore variant callers and developed a random forest classifier to filter called SNPs. Clair was the most suitable variant caller after SNP filtering. A minimum depth of 20x reads was required to confidently identify resistant determinants over the entire genome. Our findings show that metagenomic Nanopore sequencing can provide reliable diagnostic information in N. gonorrhoeae infection.

microbiology↗

bioRxiv2019

Optimizing DNA extraction methods for Nanopore sequencing of Neisseria gonorrhoeae direct from urine samples

BackgroundEmpirical gonorrhoea treatment at initial diagnosis reduces onward transmission. However, increasing resistance to multiple antibiotics may necessitate waiting for culture-based diagnostics to select an effective treatment. There is a need for same-day culture-free diagnostics that identify infection and detect antimicrobial resistance.\n\nMethodsWe investigated if Nanopore sequencing can detect sufficient N. gonorrhoeae DNA to reconstruct whole genomes directly from urine samples. We used N. gonorrhoeae spiked urine samples and samples from gonorrhoea infections to determine optimal DNA extraction methods that maximize the amount of N. gonorrhoeae DNA sequenced whilst minimizing contaminating host DNA.\n\nResultsIn simulated infections the Qiagen UCP Pathogen Mini kit provided the highest ratio N. gonorrhoeae to human DNA and the most consistent results. Depletion of human DNA with saponin increased N. gonorrhoeae yields in simulated infections, but decreased yields in clinical samples. In ten urine samples from men with symptomatic urethral gonorrhoea, [≥]87% coverage of an N. gonorrhoeae reference genome was achieved in all samples, with [≥]92% coverage breath at [≥]10-fold depth in 7 (70%) samples. In simulated infections if [≥]104 CFU/ml of N. gonorrhoeae was present, sequencing of the large majority of the genome was frequently achieved. N. gonorrhoeae could also be detected from urine in cobas PCR Media tubes and from urethral swabs, and in the presence of simulated Chlamydia co-infection.\n\nConclusionUsing Nanopore sequencing of urine samples from men with urethral gonorrhoea sufficient data can be obtained to reconstruct whole genomes in the majority of samples without the need for culture.

microbiology↗

bioRxiv2018

Panton-Valentine leukocidin is the key determinant of Staphylococcus aureus pyomyositis in a bacterial genome-wide association study

Pyomyositis is a severe bacterial infection of skeletal muscle, commonly affecting children in tropical regions and predominantly caused by Staphylococcus aureus. To understand the contribution of bacterial genomic factors to pyomyositis, we conducted a genome-wide association study of S. aureus cultured from 101 children with pyomyositis and 417 children with asymptomatic nasal carriage attending the Angkor Hospital for Children in Cambodia. We found a strong relationship between bacterial genetic variation and pyomyositis, with estimated heritability 63.8% (95% CI 49.2-78.4%). The presence of the Panton-Valentine leucocidin (PVL) locus increased the odds of pyomyositis 130-fold (p =10-17.9). The signal of association mapped both to the PVL-coding sequence and the sequence immediately upstream. Together these regions explained > 99.9% of heritability. Our results establish staphylococcal pyomyositis, like tetanus and diphtheria, as critically dependent on expression of a single toxin and demonstrate the potential for association studies to identify specific bacterial genes promoting severe human disease.

microbiology↗