Continued improvement in the development of the SARS-CoV-2 whole genome sequencing proficiency testing program.

Application of whole genome sequencing (WGS) has allowed monitoring of the emergence of variants of concern (VOC) of severe acute respiratory syndrome-related coronavirus 2 (SARS-CoV-2) globally. Genomic investigation of emerging variants and surveillance of clinical progress has reduced the public health impact of infection during the COVID-19 pandemic. These steps required developing and implementing a proficiency testing program (PTP), as WGS has been incorporated into routine reference laboratory practice.

Bioinformatic investigation of discordant sequence data for SARS-CoV-2: insights for robust genomic analysis during pandemic surveillance.

The COVID-19 pandemic has necessitated the rapid development and implementation of whole-genome sequencing (WGS) and bioinformatic methods for managing the pandemic. However, variability in methods and capabilities between laboratories has posed challenges in ensuring data accuracy. A national working group comprising 18 laboratory scientists and bioinformaticians from Australia and New Zealand was formed to improve data concordance across public health laboratories (PHLs).

Proficiency testing for SARS-CoV-2 whole genome sequencing.

Extensive studies and analyses into the molecular features of severe acute respiratory syndrome related coronavirus 2 (SARS-CoV-2) have enhanced the surveillance and investigation of its clusters and transmission worldwide. The whole genome sequencing (WGS) approach is crucial in identifying the source of infection and transmission routes by monitoring the emergence of variants over time and through communities. Varying SARS-CoV-2 genomics capacity and capability levels have been established in public health laboratories across different Australian states and territories.

Proficiency testing for SARS-CoV-2 in assuring the quality and overall performance in viral RNA detection in clinical and public health laboratories.

Diagnostic testing for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has undergone significant changes over the duration of the pandemic. In early 2020, SARS-CoV-2 specific nucleic acid testing (NAT) protocols were predominantly in-house assays developed based on protocols published in peer reviewed journals. As the pandemic has progressed, there has been an increase in the choice of testing platforms.

Proficiency testing for bacterial whole genome sequencing in assuring the quality of microbiology diagnostics in clinical and public health laboratories.

The adoption of whole genome sequencing (WGS) data over the past decade for pathogen surveillance, and decision-making for infectious diseases has rapidly transformed the landscape of clinical microbiology and public health. However, for successful transition to routine use of these techniques, it is crucial to ensure the WGS data generated meet defined quality standards for pathogen identification, typing, antimicrobial resistance detection and surveillance. Further, the ongoing development of these standards will ensure that the bioinformatic processes are capable of accurately identifying and characterising organisms of interest, and thereby facilitate the integration of WGS into routine clinical and public health laboratory setting.

Laboratory biosafety measures involving SARS-CoV-2 and the classification as a Risk Group 3 biological agent.

The current public health emergency surrounding the COVID-19 pandemic, that is the illness caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has resulted in thousands of cases in Australia since 25 January 2020 when the first case was diagnosed. This emerging virus presents particular hazards to researchers and laboratory staff in a clinical setting, highlighted by rapid and widespread global transmission. Based on the epidemiological and clinical data that have become available in mid-2020, we propose the interim classification of SARS-CoV-2 as a Risk Group 3 organism is reasonable, and discuss establishing Biosafety Level 3 (BSL-3) regulations accordingly.

Proficiency testing for the detection of Middle East respiratory syndrome coronavirus demonstrates global capacity to detect Middle East respiratory syndrome coronavirus.

The first reported case of Middle East respiratory syndrome coronavirus (MERS-CoV) infection was identified in Saudi Arabia in September 2012, since which time there have been over 2000 laboratory-confirmed cases, including 750 deaths in 27 countries. Nucleic acid testing (NAT) is the preferred method for the detection of MERS-CoV. A single round of a Proficiency Testing Program (PTP) was used to assess the capability of laboratories globally to accurately detect the presence of MERS-CoV using NAT.

Ebola Preparedness: Diagnosis Improvement Using Rapid Approaches for Proficiency Testing.

The unprecedented 2015 Ebolavirus (EBOV) outbreak in West Africa was declared a public health emergency, making diagnosis and quality of testing a global issue. The accuracy of laboratory diagnostic capacity for EBOV was assessed in 2014 to 2016 using a proficiency testing (PT) strategy developed by the Royal College of Pathologists of Australasia Quality Assurance Programs (RCPAQAP) in Biosecurity. Following a literature search, EBOV-specific gene targets were ranked according to the frequency of their use in published methods.

A simple plasmid-based system that allows rapid generation of tightly controlled gene expression in Staphylococcus aureus.

We have established a plasmid-based system that enables tightly controlled gene expression and the generation of GFP fusion proteins in Staphylococcus aureus simply and rapidly. This system takes advantage of an Escherichia coli-S. aureus shuttle vector that contains the replication region of the S.

Biochemical characterization of the multidrug regulator QacR distinguishes residues that are crucial to multidrug binding and induction of qacA transcription.

Staphylococcus aureus transcription factor QacR regulates expression of the qacA multidrug efflux determinant. In response to binding cationic lipophilic compounds, including ethidium and rhodamine 6G, QacR dissociates from the qacA operator alleviating repression. Such ligand binding uniformly induces a coil-to-helix transition of residues Thr(89)-Tyr(93) revealing an asymmetric binding pocket in QacR containing two distinct subpockets.

Mutations in TOPORS cause autosomal dominant retinitis pigmentosa with perivascular retinal pigment epithelium atrophy.

We report mutations in the gene for topoisomerase I-binding RS protein (TOPORS) in patients with autosomal dominant retinitis pigmentosa (adRP) linked to chromosome 9p21.1 (locus RP31). A positional-cloning approach, together with the use of bioinformatics, identified TOPORS (comprising three exons and encoding a protein of 1,045 aa) as the gene responsible for adRP.

Identification of suitable internal controls to study expression of a Staphylococcus aureus multidrug resistance system by quantitative real-time PCR.

Quantitative real-time PCR (qRT-PCR) has become a routine technique for gene expression analysis. Housekeeping genes are customarily used as endogenous references for the relative quantification of genes of interest. The aim of this study was to develop a quantitative real-time PCR assay to analyze gene expression in multidrug resistant Staphylococcus aureus in the presence of cationic lipophilic substrates of multidrug transport proteins.

New insights into the target site and mode of action of the antifungal protein of Aspergillus giganteus.

The antifungal protein (AFP) secreted by Aspergillus giganteus exerts growth inhibitory effects on various filamentous fungi. In order to obtain more information on the mode of action of AFP, we used transmission electron microscopy in this study to compare the cellular ultrastructure of the AFP-sensitive Aspergillus niger and of the AFP-resistant Penicillium chrysogenum upon AFP treatment. Furthermore, AFP was localized by immunogold staining in both fungi.