Biology subjects

Willcocks, S.

Publications and source records attributed to Willcocks, S..

4 recordsLinked to original sources

bioRxiv2023

Understanding Campylobacter coli isolates from the Vietnamese meat production network; a pilot study

Changing farming practices and the associated increase in the use of antibiotics are amongst the main drivers shaping the global increase of Campylobacter infections. The effects farming practices have on Campylobacter species, need to be studied at the global scale, particularly in emerging middle-income countries, where the demand for low-cost poultry meat is rising. While C. jejuni causes the majority of poultry associated diarrhoea, C. coli causes a significant amount of disease but are relatively understudied. In this study we characterised seven C. coli strains isolated from poultry farms and markets in Hanoi, Vietnam. Comprehensive data sets of bacterial Whole-Genome Sequencing; and phenotypic assays, such as, growth, motility, antimicrobial resistant testing along with virulence testing were performed to reveal the genetic relatedness and pathophysiological characteristics of seven C. coli strains. Six isolates were classified as multi-drug resistant, with all isolates resistant to ciprofloxacin, nalidixic acid and tetracycline, but susceptible to phenicols. All isolates had similar growth rates, while five were hyper-motile. Lethality of the isolates towards a tractable host-model system, larvae of the greater wax moth Galleria mellonella, often used to determine Campylobacter virulence was demonstrated for the first time for C. coli. Multilocus sequence typing data correlates with North American, European, and Asian isolates from patients suffering from gastroenteritis, emphasising the global spread of these strains. This work demonstrates that C. coli, with high levels of antimicrobial resistance, is an understudied global threat. Data summaryGenBank database with accession numbers JAKGTW000000000, JAKGTV000000000, JAKGTS000000000, JAKGTU000000000, JAKGTT000000000, JAKGTR000000000 and CP091310 https://www.ncbi.nlm.nih.gov/nuccore/JAKGTW000000000 https://www.ncbi.nlm.nih.gov/nuccore/JAKGTV000000000 https://www.ncbi.nlm.nih.gov/nuccore/JAKGTS000000000 https://www.ncbi.nlm.nih.gov/nuccore/JAKGTU000000000 https://www.ncbi.nlm.nih.gov/nuccore/JAKGTT000000000 https://www.ncbi.nlm.nih.gov/nuccore/JAKGTR000000000 https://www.ncbi.nlm.nih.gov/nuccore/CP091310.1 The authors confirm all supporting data, code and protocols have been provided within the article or through supplementary data files.

microbiology↗

bioRxiv2022

Defining the genes required for survival of Mycobacterium bovis in the bovine host offers novel insights into the genetic basis of survival of pathogenic mycobacteria

Tuberculosis has severe impacts in both humans and animals. Understanding the genetic basis of survival of both Mycobacterium tuberculosis, the human adapted species, and Mycobacterium bovis, the animal adapted species is crucial to deciphering the biology of both pathogens. There are several studies that identify the genes required for survival of M. tuberculosis in vivo using mouse models, however, there are currently no studies probing the genetic basis of survival of M. bovis in vivo. In this study we utilise transposon insertion sequencing in M. bovis to determine the genes required for survival in cattle. We identify genes encoding established mycobacterial virulence functions such as the ESX-1 secretion system, PDIM synthesis, mycobactin synthesis and cholesterol catabolism that are required in vivo. We show that, as in M. tuberculosis, phoPR is required by M. bovis in vivo despite the known defect in signalling through this system. Comparison to studies performed in glycerol adapted species such as M. bovis BCG and M. tuberculosis suggests that there are differences in the requirement for genes involved in cholesterol import (mce4 operon), oxidation (hsd) and detoxification (cyp125). We report good correlation with existing mycobacterial virulence functions, but also find several novel virulence factors, including genes involved in protein mannosylation, aspartate metabolism and glycerol-phosphate metabolism. These findings further extend our knowledge of the genetic basis of survival in vivo in bacteria that cause tuberculosis and provide insight for the development of novel diagnostics and therapeutics. ImportanceThis is the first report of the genetic requirements of an animal adapted member of the MTBC in a natural host. M. bovis has devastating impacts in cattle and bovine tuberculosis is a considerable economic, animal welfare and public health concern. The data highlight the importance of mycobacterial cholesterol catabolism and identifies several new virulence factors. Additionally, the work informs the development of novel differential diagnostics and therapeutics for TB in both human and animal populations.

microbiology↗

bioRxiv2021

Probing differences in gene essentiality between the human and animal adapted lineages of the Mycobacterium tuberculosis complex using TnSeq

Members of the Mycobacterium tuberculosis complex (MTBC) show distinct host adaptations, preferences and phenotypes despite being >99% identical at the nucleic acid level. Previous studies have explored gene expression changes between the members, however few studies have probed differences in gene essentiality. To better understand the functional impacts of the nucleic acid differences between Mycobacterium bovis and Mycobacterium tuberculosis we used the Mycomar T7 phagemid delivery system to generate whole genome transposon libraries in laboratory strains of both species and compared the essentiality status of genes during growth under identical in vitro conditions. Libraries contained insertions in 54% of possible TA sites in M. bovis and 40% of those present in M. tuberculosis, achieving similar saturation levels to those previously reported for the MTBC. The distributions of essentiality across the functional categories were similar in both species. 527 genes were found to be essential in M. bovis whereas 477 genes were essential in M. tuberculosis and 370 essential genes were common in both species. CRISPRi was successfully utilised in both species to determine the impacts of silencing genes including wag31, a gene involved in peptidoglycan synthesis and Rv2182c/Mb2204c, a gene involved in glycerophospholipid metabolism. We observed species specific differences in the response to gene silencing, with the inhibition of expression of Mb2204c in M. bovis showing significantly less growth impact than silencing its ortholog (Rv2182c) in M. tuberculosis. Given that glycerophospholipid metabolism is a validated pathway for antimicrobials, our observations suggest that target vulnerability in the animal adapted lineages cannot be assumed to be the same as the human counterpart. This is of relevance for zoonotic tuberculosis as it implies that the development of antimicrobials targeting the human adapted lineage might not necessarily be effective against the animal adapted lineage. The generation of a transposon library and the first reported utilisation of CRISPRi in M. bovis will enable the use of these tools to further probe the genetic basis of survival under disease relevant conditions.

microbiology↗

bioRxiv2021

Single Nucleotide Polymorphisms in the Bovine TLR2 Extracellular Domain Contribute to Breed and Species-Specific Innate Immune Functionality

Recent evidence suggests that several cattle breeds may be more resistant to infection with the zoonotic pathogen Mycobacterium bovis than others. Our data presented here suggests that the response to mycobacterial antigens varies in macrophages generated from Brown Swiss (BS) and Holstein Frisian (HF) cattle, two breeds belonging to the Bos taurus family. Whole genome sequencing of the Brown Swiss genome identified several potential candidate genes, in particular Toll-like Receptor-2 (TLR2) a pattern recognition receptor (PRR) that has previously been described to be involved in mycobacterial recognition. Further investigation revealed single nucleotide polymorphisms (SNP) in TLR2 that were identified between DNA isolated from cells of BS and HF cows. Interestingly, one specific SNP, H326Q, showed a different genotype frequency in two cattle subspecies, Bos taurus and Bos indicus. Cloning of the TLR2 gene and subsequent gene-reporter and chemokine assays revealed that this SNP, present in BS and Bos indicus breeds, resulted in a significantly higher response to mycobacterial antigens as well as tri-acylated lipopeptide ligands in general. Comparing wild-type and H326Q containing TLR2 responses, wild-type bovine TLR2 response showed clear, diminished mycobacterial antigen responses compared to human TLR2, however bovine TLR2 responses containing H326Q were found to be partially recovered compared to human TLR2. The creation of human:bovine TLR2 chimeras increased the response to mycobacterial antigens compared to the full-length bovine TLR2, but significantly reduced the response compared to the full-length human TLR2. Thus, our data, not only present evidence that TLR2 is a major PRR in the mammalian species-specific response to mycobacterial antigens, but furthermore, that there are clear differences between the response seen in different cattle breeds, which may contribute to their enhanced or reduced susceptibility to mycobacterial infection.

immunology↗