ACORN (A Clinically-Oriented Antimicrobial Resistance Surveillance Network) II: protocol for case based antimicrobial resistance surveillance

Background: Antimicrobial resistance surveillance is essential for empiric antibiotic prescribing, infection prevention and control policies and to drive novel antibiotic discovery. However, most existing surveillance systems are isolate-based without supporting patient-based clinical data, and not widely implemented especially in low- and middle-income countries (LMICs). Methods: A Clinically-Oriented Antimicrobial Resistance Surveillance Network (ACORN) II is a large-scale multicentre protocol which builds on the WHO Global Antimicrobial Resistance and Use Surveillance System to estimate syndromic and pathogen outcomes along with associated health economic costs. ACORN-healthcare associated infection (ACORN-HAI) is an extension study which focuses on healthcare-associated bloodstream infections and ventilator-associated pneumonia. Our main aim is to implement an efficient clinically-oriented antimicrobial resistance surveillance system, which can be incorporated as part of routine workflow in hospitals in LMICs. These surveillance systems include hospitalised patients of any age with clinically compatible acute community-acquired or healthcare-associated bacterial infection syndromes, and who were prescribed parenteral antibiotics. Diagnostic stewardship activities will be implemented to optimise microbiology culture specimen collection practices. Basic patient characteristics, clinician diagnosis, empiric treatment, infection severity and risk factors for HAI are recorded on enrolment and during 28-day follow-up. An R Shiny application can be used offline and online for merging clinical and microbiology data, and generating collated reports to inform local antibiotic stewardship and infection control policies. Discussion: ACORN II is a comprehensive antimicrobial resistance surveillance activity which advocates pragmatic implementation and prioritises improving local diagnostic and antibiotic prescribing practices through patient-centred data collection. These data can be rapidly communicated to local physicians and infection prevention and control teams. Relative ease of data collection promotes sustainability and maximises participation and scalability. With ACORN-HAI as an example, ACORN II has the capacity to accommodate extensions to investigate further specific questions of interest

Molecular epidemiology of tuberculosis in low and high incidence settings

Tuberculosis (TB) surpassed HIV/AIDS in 2015 to become the greatest infectious disease killer on the planet, while the inexorable rise of drug-resistant TB adds another layer of complexity to TB control challenges in both high and low incidence settings. The advances in molecular genotyping of Mycobacterium tuberculosis has promised new but yet validated evidence and performance indicators to inform jurisdictional TB control programs. The overall aim of this thesis was to improve our understanding of the epidemiology of M. tuberculosis through integration of traditional genotyping and geospatial methods in low and high incidence settings, exemplified, respectively, by New South Wales (NSW), Australia, and Mongolia. It also explored the added value of WGS in improving our understanding of M. tuberculosis transmission dynamics in low incidence settings. The following hypotheses were tested: The application of high-resolution genotyping of M. tuberculosis can reveal the unique features and dynamic changes in molecular epidemiology patterns over short period of time, and the synthesis of epidemiological and high-resolution M. tuberculosis genotyping data can identify spatio-temporal hotspots of potential disease transmission of relevance to TB control programs. To explore these hypotheses we examined mycobacterial interspersed repetitive unit-variable number tandem repeat (MIRU-24) genotypes of M. tuberculosis isolates recovered from patients in NSW and Mongolia, respectively, and corresponding drug susceptibility and epidemiological data. Thirty strains from NSW were also subjected to whole genome sequencing (WGS). In NSW important recent changes were demonstrated in the M. tuberculosis population structure. Three pre-dominant phylogenetic lineages (Beijing, East African Indian (EAI) and Delhi/CAS) accounted for most cases, but between 2010 and 2012, EAI strains overtook Beijing as the most prevalent genotype. This shift in M. tuberculosis population structure reflects increased migration to Australia from the Indian Sub-continent. Twenty-four loci mycobacterial interspersed repetitive unit-variable number tandem repeat (MIRU-24) demonstrated superior resolution to MIRU-12, but the resolution of Beijing lineage strains remained sub-optimal. Relatively high rates of identical MIRU-24 profiles (12.8%) suggested significant local transmission. Significant MIRU clustering of Beijing lineage strains was resolved by WGS. Genome sequencing demonstrated that only 2 strains within a 21-member Beijing cluster represented a probable recent transmission chain. The EAI clusters identified by MIRU-24 were completely resolved by WGS, demonstrating the value of WGS to limit unnecessary cluster investigation and guide better-targeted public health responses. Synthesis of geospatial and genotypic data offered important insights into the local TB transmission in NSW and indicated that the predominant majority of human cases of TB caused by Beijing lineage strains were reactivation of latent infection acquired in the country of origin rather than recent transmission. In Mongolia, programmatic management of multi-drug resistant TB (MDR-TB) led to a rapid increase in MDR-TB case finding. Spatio-temporal mapping demonstrated clustering of MDR-TB cases along the Trans-Siberian railway line, suggesting initial introduction along the railway line with subsequent spread to more remote provinces. Molecular analysis of MDR-TB isolates recovered from cases diagnosed in 2012, especially those classified as Category 1 treatment failure, revealed predominance of Beijing lineage strain, with a high percentage of MIRU-24 clustering. Combined analysis of genotypic and drug susceptibility data suggested that the MDR-TB epidemic in Mongolia has been mainly driven by Beijing lineage strains resistant to all first-line drugs. These findings have not been reported before and have significant implications for TB control efforts within Mongolia. Research objectives have been achieved and the methods and findings from the thesis could be valuable to not only to NSW and Mongolian TB control programs but could assist other countries to gain in-depth understanding of dynamics of the disease spread and to make the vision of TB free world one step closer

Demographic, clinical and MIRU-24 clustering characteristics of predominant <i>M</i>. <i>tuberculosis</i> strain lineages.

<p>Demographic, clinical and MIRU-24 clustering characteristics of predominant <i>M</i>. <i>tuberculosis</i> strain lineages.</p

Whole-genome Bayesian inference distance tree of Lineage 2 Beijing clusters with identical MIRU-24 profiles (MIRU-24 clusters).

<p>MIRU-24 cluster C is labeled in green, and MIRU cluster D in red and three SNP clusters were identified. Three libraries of the two SNP clusters were determined to represent cross-contamination during diagnostic culture are marked with an asterisk. Branch support probabilities are displayed as percentages in blue. The scale bars represents 10 substitutions per genome for the corresponding distance from a node. Parameters, output and version information for <i>mrbayes</i> can be found in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0163612#pone.0163612.s006" target="_blank">S2 File</a>.</p

Whole Genome Sequencing Demonstrates Limited Transmission within Identified <i>Mycobacterium tuberculosis</i> Clusters in New South Wales, Australia

<div><p>Australia has a low tuberculosis incidence rate with most cases occurring among recent immigrants. Given suboptimal cluster resolution achieved with 24-locus mycobacterium interspersed repetitive unit (MIRU-24) genotyping, the added value of whole genome sequencing was explored. MIRU-24 profiles of all <i>Mycobacterium tuberculosis</i> culture-confirmed tuberculosis cases diagnosed between 2009 and 2013 in New South Wales (NSW), Australia, were examined and clusters identified. The relatedness of cases within the largest MIRU-24 clusters was assessed using whole genome sequencing and phylogenetic analyses. Of 1841 culture-confirmed TB cases, 91.9% (1692/1841) had complete demographic and genotyping data. East-African Indian (474; 28.0%) and Beijing (470; 27.8%) lineage strains predominated. The overall rate of MIRU-24 clustering was 20.1% (340/1692) and was highest among Beijing lineage strains (35.7%; 168/470). One Beijing and three East-African Indian (EAI) clonal complexes were responsible for the majority of observed clusters. Whole genome sequencing of the 4 largest clusters (30 isolates) demonstrated diverse single nucleotide polymorphisms (SNPs) within identified clusters. All sequenced EAI strains and 70% of Beijing lineage strains clustered by MIRU-24 typing demonstrated distinct SNP profiles. The superior resolution provided by whole genome sequencing demonstrated limited <i>M</i>. <i>tuberculosis</i> transmission within NSW, even within identified MIRU-24 clusters. Routine whole genome sequencing could provide valuable public health guidance in low burden settings.</p></div

Tuberculosis incidence and genotypic clustering rate in NSW, Australia.

<p>MIRU-24–24-locus mycobacterium interspersed repetitive unit strain typing method.</p

Whole-genome Bayesian inference distance tree of Lineage 1 East African Indian (EAI) clusters with identical MIRU-24 profiles (MIRU-24 clusters).

<p>MIRU-24 cluster A is labeled in red, and MIRU-24 cluster B in green and were only distantly related (>100 SNP differences) on whole genome sequencing without any SNP clusters identified.</p