Biochemistry and Pharmacology - Research Publications

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    Trajectories of childhood immune development and respiratory health relevant to asthma and allergy.
    Tang, HH ; Teo, SM ; Belgrave, DC ; Evans, MD ; Jackson, DJ ; Brozynska, M ; Kusel, MM ; Johnston, SL ; Gern, JE ; Lemanske, RF ; Simpson, A ; Custovic, A ; Sly, PD ; Holt, PG ; Holt, KE ; Inouye, M (eLife Sciences Publications, Ltd, 2018-10-15)
    Events in early life contribute to subsequent risk of asthma; however, the causes and trajectories of childhood wheeze are heterogeneous and do not always result in asthma. Similarly, not all atopic individuals develop wheeze, and vice versa. The reasons for these differences are unclear. Using unsupervised model-based cluster analysis, we identified latent clusters within a prospective birth cohort with deep immunological and respiratory phenotyping. We characterised each cluster in terms of immunological profile and disease risk, and replicated our results in external cohorts from the UK and USA. We discovered three distinct trajectories, one of which is a high-risk 'atopic' cluster with increased propensity for allergic diseases throughout childhood. Atopy contributes varyingly to later wheeze depending on cluster membership. Our findings demonstrate the utility of unsupervised analysis in elucidating heterogeneity in asthma pathogenesis and provide a foundation for improving management and prevention of childhood asthma.
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    FastSpar: rapid and scalable correlation estimation for compositional data
    Watts, SC ; Ritchie, SC ; Inouye, M ; Holt, KE ; Stegle, O (OXFORD UNIV PRESS, 2019-03-15)
    A common goal of microbiome studies is the elucidation of community composition and member interactions using counts of taxonomic units extracted from sequence data. Inference of interaction networks from sparse and compositional data requires specialized statistical approaches. A popular solution is SparCC, however its performance limits the calculation of interaction networks for very high-dimensional datasets. Here we introduce FastSpar, an efficient and parallelizable implementation of the SparCC algorithm which rapidly infers correlation networks and calculates P-values using an unbiased estimator. We further demonstrate that FastSpar reduces network inference wall time by 2–3 orders of magnitude compared to SparCC.
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    Airway Microbiota Dynamics Uncover a Critical Window for Interplay of Pathogenic Bacteria and Allergy in Childhood Respiratory Disease
    Teo, SM ; Tang, HHF ; Mok, D ; Judd, LM ; Watts, SC ; Pham, K ; Holt, BJ ; Kusel, M ; Serralha, M ; Troy, N ; Bochkov, YA ; Grindle, K ; Lemanske, RF ; Johnston, SL ; Gern, JE ; Sly, PD ; Holt, PG ; Holt, KE ; Inouye, M (CELL PRESS, 2018-09-12)
    Repeated cycles of infection-associated lower airway inflammation drive the pathogenesis of persistent wheezing disease in children. In this study, the occurrence of acute respiratory tract illnesses (ARIs) and the nasopharyngeal microbiome (NPM) were characterized in 244 infants through their first five years of life. Through this analysis, we demonstrate that >80% of infectious events involve viral pathogens, but are accompanied by a shift in the NPM toward dominance by a small range of pathogenic bacterial genera. Unexpectedly, this change frequently precedes the detection of viral pathogens and acute symptoms. Colonization of illness-associated bacteria coupled with early allergic sensitization is associated with persistent wheeze in school-aged children, which is the hallmark of the asthma phenotype. In contrast, these bacterial genera are associated with "transient wheeze" that resolves after age 3 years in non-sensitized children. Thus, to complement early allergic sensitization, monitoring NPM composition may enable early detection and intervention in high-risk children.
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    The Infant Nasopharyngeal Microbiome Impacts Severity of Lower Respiratory Infection and Risk of Asthma Development
    Teo, SM ; Mok, D ; Pham, K ; Kusel, M ; Serralha, M ; Troy, N ; Holt, BJ ; Hales, BJ ; Walker, ML ; Hollams, E ; Bochkov, YA ; Grindle, K ; Johnston, SL ; Gern, JE ; Sly, PD ; Holt, PG ; Holt, KE ; Inouye, M (CELL PRESS, 2015-05-13)
    The nasopharynx (NP) is a reservoir for microbes associated with acute respiratory infections (ARIs). Lung inflammation resulting from ARIs during infancy is linked to asthma development. We examined the NP microbiome during the critical first year of life in a prospective cohort of 234 children, capturing both the viral and bacterial communities and documenting all incidents of ARIs. Most infants were initially colonized with Staphylococcus or Corynebacterium before stable colonization with Alloiococcus or Moraxella. Transient incursions of Streptococcus, Moraxella, or Haemophilus marked virus-associated ARIs. Our data identify the NP microbiome as a determinant for infection spread to the lower airways, severity of accompanying inflammatory symptoms, and risk for future asthma development. Early asymptomatic colonization with Streptococcus was a strong asthma predictor, and antibiotic usage disrupted asymptomatic colonization patterns. In the absence of effective anti-viral therapies, targeting pathogenic bacteria within the NP microbiome could represent a prophylactic approach to asthma.
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    SRST2: Rapid genomic surveillance for public health and hospital microbiology labs
    Inouye, M ; Dashnow, H ; Raven, L-A ; Schultz, MB ; Pope, BJ ; Tomita, T ; Zobel, J ; Holt, KE (BMC, 2014-11-20)
    Rapid molecular typing of bacterial pathogens is critical for public health epidemiology, surveillance and infection control, yet routine use of whole genome sequencing (WGS) for these purposes poses significant challenges. Here we present SRST2, a read mapping-based tool for fast and accurate detection of genes, alleles and multi-locus sequence types (MLST) from WGS data. Using >900 genomes from common pathogens, we show SRST2 is highly accurate and outperforms assembly-based methods in terms of both gene detection and allele assignment. We include validation of SRST2 within a public health laboratory, and demonstrate its use for microbial genome surveillance in the hospital setting. In the face of rising threats of antimicrobial resistance and emerging virulence among bacterial pathogens, SRST2 represents a powerful tool for rapidly extracting clinically useful information from raw WGS data. Source code is available from http://katholt.github.io/srst2/.
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    In silico serotyping of E. coli from short read data identifies limited novel O-loci but extensive diversity of O:H serotype combinations within and between pathogenic lineages
    Ingle, DJ ; Valcanis, M ; Kuzevski, A ; Tauschek, M ; Inouye, M ; Stinear, T ; Levine, MM ; Robins-Browne, RM ; Holt, KE (MICROBIOLOGY SOC, 2016-07-01)
    The lipopolysaccharide (O) and flagellar (H) surface antigens of Escherichia coli are targets for serotyping that have traditionally been used to identify pathogenic lineages. These surface antigens are important for the survival of E. coli within mammalian hosts. However, traditional serotyping has several limitations, and public health reference laboratories are increasingly moving towards whole genome sequencing (WGS) to characterize bacterial isolates. Here we present a method to rapidly and accurately serotype E. coli isolates from raw, short read WGS data. Our approach bypasses the need for de novo genome assembly by directly screening WGS reads against a curated database of alleles linked to known and novel E. coli O-groups and H-types (the EcOH database) using the software package srst2. We validated the approach by comparing in silico results for 197 enteropathogenic E. coli isolates with those obtained by serological phenotyping in an independent laboratory. We then demonstrated the utility of our method to characterize isolates in public health and clinical settings, and to explore the genetic diversity of >1500 E. coli genomes from multiple sources. Importantly, we showed that transfer of O- and H-antigen loci between E. coli chromosomal backbones is common, with little evidence of constraints by host or pathotype, suggesting that E. coli 'strain space' may be virtually unlimited, even within specific pathotypes. Our findings show that serotyping is most useful when used in combination with strain genotyping to characterize microevolution events within an inferred population structure.
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    Developmental patterns in the nasopharyngeal microbiome during infancy are associated with asthma risk
    Tang, HHF ; Lang, A ; Teo, SM ; Judd, LM ; Gangnon, R ; Evans, MD ; Lee, KE ; Vrtis, R ; Holt, PG ; Lemanske, RF ; Jackson, DJ ; Holt, KE ; Inouye, M ; Gern, JE (MOSBY-ELSEVIER, 2021-05-05)
    BACKGROUND: Studies indicate that the nasal microbiome may correlate strongly with the presence or future risk of childhood asthma. OBJECTIVES: In this study, we tested whether developmental trajectories of the nasopharyngeal microbiome in early life and the composition of the microbiome during illnesses were related to risk of childhood asthma. METHODS: Children participating in the Childhood Origins of Asthma study (N = 285) provided nasopharyngeal mucus samples in the first 2 years of life, during routine healthy study visits (at 2, 4, 6, 9, 12, 18, and 24 months of age), and during episodes of respiratory illnesses, all of which were analyzed for respiratory viruses and bacteria. We identified developmental trajectories of early-life microbiome composition, as well as predominant bacteria during respiratory illnesses, and we correlated these with presence of asthma at 6, 8, 11, 13, and 18 years of age. RESULTS: Of the 4 microbiome trajectories identified, a Staphylococcus-dominant microbiome in the first 6 months of life was associated with increased risk of recurrent wheezing by age 3 years and asthma that persisted throughout childhood. In addition, this trajectory was associated with the early onset of allergic sensitization. During wheezing illnesses, detection of rhinoviruses and predominance of Moraxella were associated with asthma that persisted throughout later childhood. CONCLUSION: In infancy, the developmental composition of the microbiome during healthy periods and the predominant microbes during acute wheezing illnesses are both associated with the subsequent risk of developing persistent childhood asthma.
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    GeneMates: an R package for detecting horizontal gene co-transfer between bacteria using gene-gene associations controlled for population structure
    Wan, Y ; Wick, RR ; Zobel, J ; Ingle, DJ ; Inouye, M ; Holt, KE (BMC, 2020-09-24)
    BACKGROUND: Horizontal gene transfer contributes to bacterial evolution through mobilising genes across various taxonomical boundaries. It is frequently mediated by mobile genetic elements (MGEs), which may capture, maintain, and rearrange mobile genes and co-mobilise them between bacteria, causing horizontal gene co-transfer (HGcoT). This physical linkage between mobile genes poses a great threat to public health as it facilitates dissemination and co-selection of clinically important genes amongst bacteria. Although rapid accumulation of bacterial whole-genome sequencing data since the 2000s enables study of HGcoT at the population level, results based on genetic co-occurrence counts and simple association tests are usually confounded by bacterial population structure when sampled bacteria belong to the same species, leading to spurious conclusions. RESULTS: We have developed a network approach to explore WGS data for evidence of intraspecies HGcoT and have implemented it in R package GeneMates ( github.com/wanyuac/GeneMates ). The package takes as input an allelic presence-absence matrix of interested genes and a matrix of core-genome single-nucleotide polymorphisms, performs association tests with linear mixed models controlled for population structure, produces a network of significantly associated alleles, and identifies clusters within the network as plausible co-transferred alleles. GeneMates users may choose to score consistency of allelic physical distances measured in genome assemblies using a novel approach we have developed and overlay scores to the network for further evidence of HGcoT. Validation studies of GeneMates on known acquired antimicrobial resistance genes in Escherichia coli and Salmonella Typhimurium show advantages of our network approach over simple association analysis: (1) distinguishing between allelic co-occurrence driven by HGcoT and that driven by clonal reproduction, (2) evaluating effects of population structure on allelic co-occurrence, and (3) direct links between allele clusters in the network and MGEs when physical distances are incorporated. CONCLUSION: GeneMates offers an effective approach to detection of intraspecies HGcoT using WGS data.
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    Neonatal genetics of gene expression reveal potential origins of autoimmune and allergic disease risk
    Huang, QQ ; Tang, HHF ; Teo, SM ; Mok, D ; Ritchie, SC ; Nath, AP ; Brozynska, M ; Salim, A ; Bakshi, A ; Holt, BJ ; Khor, CC ; Sly, PD ; Holt, PG ; Holt, KE ; Inouye, M (NATURE PORTFOLIO, 2020-07-28)
    Chronic immune-mediated diseases of adulthood often originate in early childhood. To investigate genetic associations between neonatal immunity and disease, we map expression quantitative trait loci (eQTLs) in resting myeloid cells and CD4+ T cells from cord blood samples, as well as in response to lipopolysaccharide (LPS) or phytohemagglutinin (PHA) stimulation, respectively. Cis-eQTLs are largely specific to cell type or stimulation, and 31% and 52% of genes with cis-eQTLs have response eQTLs (reQTLs) in myeloid cells and T cells, respectively. We identified cis regulatory factors acting as mediators of trans effects. There is extensive colocalisation between condition-specific neonatal cis-eQTLs and variants associated with immune-mediated diseases, in particular CTSH had widespread colocalisation across diseases. Mendelian randomisation shows causal neonatal gene expression effects on disease risk for BTN3A2, HLA-C and others. Our study elucidates the genetics of gene expression in neonatal immune cells, and aetiological origins of autoimmune and allergic diseases.
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    Frequent transmission of the Mycobacterium tuberculosis Beijing lineage and positive selection for the EsxW Beijing variant in Vietnam
    Holt, KE ; McAdam, P ; Phan, VKT ; Nguyen, TTT ; Dang, TMH ; Nguyen, NL ; Nguyen, HL ; Nguyen, TQN ; Hoang, TH ; Vu, TNH ; Thwaites, G ; Edwards, DJ ; Nath, AP ; Pham, K ; Ascher, DB ; Farrar, J ; Khor, CC ; Teo, YY ; Inouye, M ; Caws, M ; Dunstan, SJ (NATURE PUBLISHING GROUP, 2018-06-01)
    To examine the transmission dynamics of Mycobacterium tuberculosis (Mtb) isolated from tuberculosis patients in Ho Chi Minh City, Vietnam, we sequenced the whole genomes of 1,635 isolates and compared these with 3,144 isolates from elsewhere. The data identify an underlying burden of disease caused by the endemic Mtb lineage 1 associated with the activation of long-term latent infection, and a threefold higher burden associated with the more recently introduced Beijing lineage and lineage 4 Mtb strains. We find that Beijing lineage Mtb is frequently transferred between Vietnam and other countries, and detect higher levels of transmission of Beijing lineage strains within this host population than the endemic lineage 1 Mtb. Screening for parallel evolution of Beijing lineage-associated SNPs in other Mtb lineages as a signal of positive selection, we identify an alteration in the ESX-5 type VII-secreted protein EsxW, which could potentially contribute to the enhanced transmission of Beijing lineage Mtb in Vietnamese and other host populations.