External Quality Assessment (EQA) for SARS-CoV-2 RNA Point-of-Care Testing in Primary Healthcare Services: Analytical Performance over Seven EQA Cycles.

In April 2020, the Aboriginal and Torres Strait Islander COVID-19 Point-of-Care (POC) Testing Program was initiated to improve access to rapid molecular-based SARS-CoV-2 detection in First Nations communities. At capacity, the program reached 105 health services across Australia. An external review estimated the program contributed to averting between 23,000 and 122,000 COVID-19 infections within 40 days of the first infection in a remote community, equating to cost savings of between AU$337 million and AU$1.8 billion. Essential to the quality management of this program, a customised External Quality Assessment (EQA) program was developed with the Royal College of Pathologists of Australasia Quality Assurance Programs (RCPAQAP). From July 2020 to May 2022, SARS-CoV-2 EQA participation ranged from 93 to 100%. Overall concordance of valid EQA results was high (98%), with improved performance following the first survey. These results are consistent with those reported by 12 Australian and 4 New Zealand laboratories for three SARS-CoV-2 RNA EQA surveys in March 2020, demonstrating that SARS-CoV-2 RNA POC testing in primary care settings can be performed to an equivalent laboratory analytical standard. More broadly, this study highlights the value of quality management practices in real-world testing environments and the benefits of ongoing EQA program participation.

Molecular Detection of Neglected Tropical Diseases: The Case for Automated Near-Point-of-Care Diagnosis of Leishmaniasis.

Neglected tropical diseases affect those in poorer nations disproportionately across the globe. One example of these, leishmaniasis, is a debilitating and potentially fatal parasitic infection. Molecular detection of this disease can provide accurate and fast diagnosis, and with near point-of-care technologies, detection can be provided in many health-care settings. Traditionally, the perceived limitations to such detection methods have hindered their provision to resource-limited nations, but new technologies and techniques are helping to overcome these perceptions. The current pandemic offers an opportunity to maintain and develop further advances, ensuring molecular diagnostics are accessible to all.

Laboratory diagnostics for human Leishmania infections: a polymerase chain reaction-focussed review of detection and identification methods.

Leishmania infections span a range of clinical syndromes and impact humans from many geographic foci, but primarily the world's poorest regions. Transmitted by the bite of a female sand fly, Leishmania infections are increasing with human movement (due to international travel and war) as well as with shifts in vector habitat (due to climate change). Accurate diagnosis of the 20 or so species of Leishmania that infect humans can lead to the successful treatment of infections and, importantly, their prevention through modelling and intervention programs. A multitude of laboratory techniques for the detection of Leishmania have been developed over the past few decades, and although many have drawbacks, several of them show promise, particularly molecular methods like polymerase chain reaction. This review provides an overview of the methods available to diagnostic laboratories, from traditional techniques to the now-preferred molecular techniques, with an emphasis on polymerase chain reaction-based detection and typing methods.

Semi-Quantitative, Duplexed qPCR Assay for the Detection of Leishmania spp. Using Bisulphite Conversion Technology.

Leishmaniasis is caused by the flagellated protozoan Leishmania, and is a neglected tropical disease (NTD), as defined by the World Health Organisation (WHO). Bisulphite conversion technology converts all genomic material to a simplified form during the lysis step of the nucleic acid extraction process, and increases the efficiency of multiplex quantitative polymerase chain reaction (qPCR) reactions. Through utilization of qPCR real-time probes, in conjunction with bisulphite conversion, a new duplex assay targeting the 18S rDNA gene region was designed to detect all Leishmania species. The assay was validated against previously extracted DNA, from seven quantitated DNA and cell standards for pan-Leishmania analytical sensitivity data, and 67 cutaneous clinical samples for cutaneous clinical sensitivity data. Specificity was evaluated by testing 76 negative clinical samples and 43 bacterial, viral, protozoan and fungal species. The assay was also trialed in a side-by-side experiment against a conventional PCR (cPCR), based on the Internal transcribed spacer region 1 (ITS1 region). Ninety-seven percent of specimens from patients that previously tested positive for Leishmania were positive for Leishmania spp. with the bisulphite conversion assay, and a limit of detection (LOD) of 10 copies per PCR was achieved, while the LOD of the ITS1 methodology was 10 cells/1000 genomic copies per PCR. This method of rapid, accurate and simple detection of Leishmania can lead to improved diagnosis, treatment and public health outcomes.