Biology subjects
Publications and source records attributed to Huws, S. A..
Milk is a challenging matrix to extract sufficient microbial DNA from for downstream analysis. This study assessed fourteen DNA extraction protocols for their DNA outputs. An adaptation of the QIAGEN DNeasy PowerSoil kit, which increased initial sample volume and maintained all volume of lysate following bead beating proved most effective.
BackgroundThe reserves of rock phosphate, used to produce fertilisers, are being exhausted by the ever-increasing pressure to meet the food demands of a growing world population. Sustainable alternatives are urgently needed, and struvite, a renewable phosphorus fertiliser recovered from waste streams such as animal manure, offers a promising solution. However, other manure-derived fertilisers have been associated with the promotion of antimicrobial resistance (AMR) in soil, raising concerns about whether struvite might pose similar risks. Despite this, the impact of struvite on the soil resistome remains poorly understood. MethodsThis study assesses the AMR risk associated with Xiamen struvite (XIA), produced from piggery wastewater, compared with two commercial fertilisers: Crystal Green (CG) and Triple Super Phosphate (TSP), and an untreated control soil. Following a three-month fertilisation period, rhizospheric soils were analysed using mass spectrometry to investigate antimicrobial residues in the soil, and shotgun metagenomics and culture-based assays, and bacterial isolate characterisation to identify bacterial AMR linked to the treatments. Notably, this is the first study to apply culturing methods to investigate struvites impact on AMR. Results and conclusionsWhile XIA struvite soil fertilisation led to an increase in total culturable bacterial load, (CFU/g), there was no evidence of antimicrobial residues detected in soil, as confirmed by mass spectrometry. Metagenomic profiling further revealed no significant alterations to antimicrobial resistance genes (ARGs) across all treatments, including CG, TSP, and untreated controls. Culture-based analysis supported these findings, with tetracycline-resistant bacteria undetected and only minimal resistance observed to erythromycin, florfenicol, and neomycin. Although resistance levels to amoxicillin, streptomycin, and trimethoprim were higher in XIA-treated soils, this appeared to reflect a broader increase in microbial load rather than selective enrichment of resistant strains. These data suggest that piggery-derived struvite does not impose a clear AMR risk in soil and performs similarly to conventional fertilisers. While it appears to increase total bacterial load, there is no evidence of selective enrichment for AMR bacteria. However, variability in antimicrobial and ARG content across studies highlights the need for standardised production specifications for struvites to ensure safety and consistency. This study provides the most comprehensive assessment to date of struvites AMR implications, supporting its potential as a safe, sustainable fertiliser and contributing valuable insights to inform regulatory guidance.
Ruminant products are consumed widely on a global level due to their high protein and micronutrient density. However, ruminant production is a major source of greenhouse gas emissions, particularly with respect to methane (CH), with ruminants contributing 33% of all anthropogenic CH emissions. CH is produced due to the natural fermentative processes undertaken by the complex rumen microbiome, primarily via the utilisation of hydrogen by rumen archaea to form CH. Previous studies have shown that feeding the red seaweed Asparagopsis taxiformis (ASP) to ruminants can reduce CHemissions from beef cattle by up to 80% (Roque et al., 2021). Nevertheless, the mechanism of action of this seaweed in terms of effects on the rumen microbiome is largely unknown, which is the main focus of this study. Six Nordic Red cows at 122 {+/-} 13.7 (mean {+/-} SD) days in milk were divided into 3 blocks by milk yield in Latin square design and fed grass silage and a commercial concentrate (60:40) either with or without 0.5% ASP on an organic matter basis. Rumen fluid was collected 19 days into each experimental period, with a holistic approach using both assembly and read mapping based approaches to interrogate taxonomic, functional, and ecological shifts applied to metagenomic data. We show that ASP reduces methane production not only through direct inhibition of methanogens but also by disrupting cobamide-dependent metabolic pathways and redirecting carbon flow toward pyruvate and propionate rather than acetate and methane. For the first time, specific enzymes involved in vitamin B12 (cobamide) biosynthesis are identified as suppressed by ASP, and microbial taxa contributing to these functional changes are elucidated showing both niche displacement and resilience within the rumen microbiome. These findings offer new mechanistic insight into how red seaweed supplementation modulates the rumen microbiome, supporting its potential role in sustainable ruminant production.
Antimicrobial Peptides (AMPs) are immune effectors that are key components of the invertebrate innate immune system providing protection against pathogenic microbes. Parasitic helminths share complex interactions with their hosts and closely associated microbiota that are likely regulated by a diverse portfolio of antimicrobial immune effectors including AMPs. Knowledge of helminth AMPs has largely been derived from nematodes, whereas the flatworm AMP repertoire has not been described. This study highlights limitations in the homology-based approaches, used to identify putative nematode AMPs, for the characterisation of flatworm AMPs, and reveals that innovative algorithmic AMP prediction approaches provide an alternative strategy for novel helminth AMP discovery. The data presented here: (i) reveal that flatworms do not encode traditional lophotrochozoan AMP groups (Big Defensin, CS{beta} peptides and Myticalin); (ii) describe a unique integrated computational pipeline for the discovery of novel helminth AMPs; (iii) reveal >16,000 putative AMP-like peptides across 127 helminth species; (iv) highlight that cysteine-rich peptides dominate helminth AMP-like peptide profiles; (v) uncover eight novel helminth AMP-like peptides with diverse antibacterial activities, and (vi) demonstrate the detection of AMP-like peptides from helminth biofluids. These data represent a significant advance in our understanding of the putative helminth AMP repertoire and underscore a potential untapped source of antimicrobial diversity which may provide opportunities for the discovery of novel antimicrobials. Further, unravelling the role of endogenous worm-derived antimicrobials and their potential to influence host-worm-microbiome interactions may be exploited for the development of unique helminth control approaches. Author summaryInvertebrate antimicrobial peptides (AMPs) form the first line of defence against pathogenic microbes. Helminths are worms (flatworm, roundworm) that live in microbe-rich environments throughout their lifecycles however little is known about how they protect themselves against pathogens or how they interact with microbes. Understanding AMP profiles in helminths, their importance to helminth biology, and how they shape microbial communities could reveal novel approaches for anthelmintic and/or antimicrobial development. In this study we describe a novel integrated homology- and computational-based pipeline for the discovery of helminth AMPs. This approach revealed that, whilst flatworms do not possess traditional AMPs, they have a repertoire of unique AMP-like peptides that are predominantly cysteine-rich. Significantly eight novel helminth AMP-like peptides, discovered using this pipeline, have antibacterial activities against a range of bacteria highlighting their potential as novel antimicrobials. Further, peptidomics analyses demonstrate the presence of AMP-like peptides in helminth body fluids supporting the need to further characterise these peptides and their function(s) in helminths. These data present novel opportunities to better understand helminth biology, discover new antimicrobials and develop future control strategies for helminth parasites.
Antimicrobial Peptides (AMPs) are key constituents of the invertebrate innate immune system where they provide critical protection against microbial threat. Knowledge of AMP complements within phylum Nematoda is limited however nematodes adopt diverse life strategies and frequently reside in microbe-rich environments such that they are likely possess broad AMP profiles with bioactivity against a range of microbiota. Indeed, parasitic nematode AMPs likely have roles in defence against invading pathogens and modulation of the host microbiome. In this study the distribution and abundance of AMP-encoding genes were examined in 134 nematode genomes providing the most comprehensive profile of AMPs within phylum Nematoda. We reveal that phylum Nematoda is AMP-rich and -diverse, where 5887 genes encode AMPs. Genome and transcriptome analyses broadly reveal: (i) AMP family profiles that are influenced by nematode lifestyle where free-living nematodes appear to have an expansion of AMPs relative to parasitic species; (ii) major differences in the AMP profiles between nematode clades where Clade 9/V and 10/IV species possess expanded AMP repertoires; (iii) AMP families with highly restricted profiles (e.g. Cecropins and Diapausins) and others [e.g. Nemapores and Glycine Rich Secreted Peptides (GRSPs)] which are more widely distributed; (iv) complexity in the distribution and abundance of Defensin subfamily members; and (v) expression of AMPs in key nematode life stages. These data indicate that phylum Nematoda has a diverse array of AMPs and underscores the need to interrogate AMP function to unravel their importance to nematode biology and host-worm-microbiome interactions. Enhanced understanding of the Nematode Antimicrobial Peptidome will inform drug discovery pipelines for pathogen control.
Honey bees use plant material to manufacture their own food. These insect pollinators visit flowers repeatedly to collect nectar and pollen, which are shared with other hive bees to produce honey and beebread. While producing these products, beehives accumulate a tremendous amount of microbes, including bacteria that derive from plants and different parts of the honey bees body. In this study, we conducted 16S rDNA metataxonomic analysis on honey and beebread samples that were collected from 15 beehives in the southeast of England in order to quantify the bacteria associated with beehives. The results highlighted that honeybee products carry a significant variety of bacterial groups that comprise bee commensals, environmental bacteria and pathogens of plants and animals. Remarkably, this bacterial diversity differs amongst the beehives, suggesting a defined fingerprint that is affected, not only by the nectar and pollen gathered from local plants, but also from other environmental sources. In summary, our results show that every hive possesses their own distinct microbiome, and that honeybee products are valuable indicators of the bacteria present in the beehives and their surrounding environment.
Herein we report the identification and characterisation of two linear antimicrobial peptides (AMPs), HG2 and HG4, with activity against a wide range of multidrug resistant (MDR) bacteria, especially methicillin resistant Staphylococcus aureus (MRSA) strains, a highly problematic group of Gram-positive bacteria in the hospital and community environment. To identify the novel AMPs presented here, we employed the classifier model design, a feature extraction method using molecular descriptors for amino acids for the analysis, visualization, and interpretation of AMP activities from a rumen metagenomic dataset. This allowed for the in silico discrimination of active and inactive peptides in order to define a small number of promising novel lead AMP test candidates for chemical synthesis and experimental evaluation. In vitro data suggest that the chosen AMPs are fast acting, show strong biofilm inhibition and dispersal activity and are efficacious in an in vivo model of MRSA USA300 infection, whilst showing little toxicity to human erythrocytes and human primary cell lines ex vivo. Observations from biophysical AMP-lipid-interactions and electron microscopy suggest that the newly identified peptides interact with the cell membrane and may be involved in the inhibition of other cellular processes. Amphiphilic conformations associated with membrane disruption are also observed in 3D molecular modelling of the peptides. HG2 and HG4 both preferentially bind to MRSA total lipids rather than with human cell lipids indicating that HG4 may form superior templates for safer therapeutic candidates for MDR bacterial infections.\n\nAuthor SummaryWe are losing our ability to treat multidrug resistant (MDR) bacteria, otherwise known as superbugs. This poses a serious global threat to human health as bacteria are increasingly acquiring resistance to antibiotics. There is therefore urgent need to intensify our efforts to develop new safer alternative drug candidates. We emphasise the usefulness of complementing wet-lab and in silico techniques for the rapid identification of new drug candidates from environmental samples, especially antimicrobial peptides (AMPs). HG2 and HG4, the AMPs identified in our study show promise as effective therapies for the treatment of methicillin resistant Staphylococcus aureus infections both in vitro and in vivo whilst having little cytotoxicity against human primary cells, a step forward in the fight against MDR infections.
Antimicrobial peptides (AMPs) are promising drug candidates to target multi-drug resistant bacteria. The rumen microbiome presents an underexplored resource for the discovery of novel microbial enzymes and metabolites, including AMPs. Using functional screening and computational approaches, we identified 181 potentially novel AMPs from a rumen bacterial metagenome. Here, we show that three of the selected AMPs (Lynronne-1, 2 and 3) were effective against numerous bacterial pathogens, including methicillin resistant Staphylococcus aureus (MRSA). No decrease in MRSA susceptibility was observed after 25 days of sub-lethal exposure to these AMPs. The AMPs bound preferentially to bacterial membrane lipids and induced membrane permeability leading to cytoplasmic leakage. Topical administration of Lynronne-1 (10% w/v) to a mouse model of MRSA wound infection elicited a significant reduction in bacterial counts, which was comparable to treatment with 2% mupirocin ointment. Our findings indicate that the rumen microbiome may provide viable alternative antimicrobials for future therapeutic application.