In 2015, a marked increase in vancomycin-resistant Enterococcus faecium (VREfm) isolation was detected at the Royal Hobart Hospital, Australia. The primary objective of this work was to examine the dynamics of VREfm transmission using whole genome data mapped to public health surveillance information. Screening and clinical isolates of VREfm from patients were typed for the specific vancomycin-resistance locus present. Of total isolates collected from 2014–2016 (n = 222), 15.3% and 84.7% harboured either the vanA or the vanB vancomycin-resistance locus, respectively. Whole-genome sequencing of 80 isolates was performed in conjunction with single-nucleotide polymorphic (SNP) analysis and in silico multi-locus sequence typing (MLST). Among the isolates sequenced, 5 phylogenetic clades were identified. The largest vanB clade belonged to MLST sequence type ST796 and contained clinical isolates from VREfm infections that clustered closely with isolates from colonised patients. Correlation of VREfm genotypes with spatio-temporal patient movements detected potential points of transmission within the hospital. ST80 emerged as the major vanA sequence type for which the most likely index case of a patient cluster was ascertained from SNP analyses. This work has identified the dominant clones associated with increased VREfm prevalence in a healthcare setting, and their likely direction of transmission.

Nontypeable Haemophilus influenzae (NTHi) colonizes human upper respiratory airways and plays a key role in the course and pathogenesis of acute exacerbations of chronic obstructive pulmonary disease (COPD). Currently, it is not possible to distinguish COPD isolates of NTHi from other clinical isolates of NTHi using conventional genotyping methods. Here, we analysed the core and accessory genome of 568 NTHi isolates, including 40 newly sequenced isolates, to look for genetic distinctions between NTHi isolates from COPD with respect to other illnesses, including otitis media, meningitis and pneumonia. Phylogenies based on polymorphic sites in the core-genome did not show discrimination between NTHi strains collected from different clinical phenotypes. However, pan-genome-wide association studies identified 79 unique NTHi accessory genes that were significantly associated with COPD. Furthermore, many of the COPD-related NTHi genes have known or predicted roles in virulence, transmembrane transport of metal ions and nutrients, cellular respiration and maintenance of redox homeostasis. This indicates that specific genes may be required by NTHi for its survival or virulence in the COPD lung. These results advance our understanding of the pathogenesis of NTHi infection in COPD lungs.

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Bacterial whole genome sequencing (WGS) is becoming a widely-used technique in research, clinical diagnostic, and public health laboratories. It enables high resolution characterization of bacterial pathogens in terms of properties that include antibiotic resistance, molecular epidemiology, and virulence. The introduction of next-generation sequencing instrumentation has made WGS attainable in terms of costs. However, the lack of a beginner’s protocol for WGS still represents a barrier to its adoption in some settings. Here, we present detailed step-by-step methods for obtaining WGS data from a range of different bacteria (Gram-positive, Gram-negative, and acid-fast) using the Illumina platform. Modifications have been performed with respect to DNA extraction and library normalization to maximize the output from the laboratory consumables invested. The protocol represents a simplified and reproducible method for producing high quality sequencing data. The key advantages of this protocol include: simplicity of the protocol for users with no prior genome sequencing experience and reproducibility of the protocol across a wide range of bacteria.

Inter-Institutional Analyses of VRE in Tasmania smaller hospitals, the North West Regional Hospital (NRH) and the Mersey Community Hospital (MCH) found that ST1421 was present in both of these institutions in 2017. These findings illustrate the spread of a recently described sequence type of VREfm, ST1421, to multiple hospitals in an Australian state within a relatively short time span.