Gemma Robertson

Genetic analysis of Giardia and Cryptosporidium from people in Northern Australia using PCR-based tools ☆

To date, there has been limited genetic study of the gastrointestinal pathogens Giardia and Cryptosporidium in northern parts of Australia. Here, PCR-based methods were used for the genetic characterization of Giardia and Cryptosporidium from 695 people with histories of gastrointestinal disorders from the tropical North of Australia. Genomic DNAs from fecal samples were subjected to PCR-based analyses of regions from the triose phosphate isomerase (tpi), small subunit (SSU) of the nuclear ribosomal RNA and/or the glycoprotein (gp60) genes. Giardia and Cryptosporidium were detected in 13 and four of the 695 samples, respectively. Giardia duodenalis assemblages A and B were found in 4 (31%) and 9 (69%) of the 13 samples in persons of <9 years of age. Cryptosporidium hominis (subgenotype IdA18), Cryptosporidium mink genotype (subgenotype IIA16R1) and C. felis were also identified in single patients of 11-21 years of age. Future studies might focus on a comparative study of these and other protists in rural communities in Northern Australia.

Rapid diagnostics for melioidosis: a comparative study of a novel lateral flow antigen detection assay

The rapid diagnosis of septicaemic melioidosis will have an impact on reduction of mortality. Currently, this relies almost exclusively upon culture of the causative agent Burkholderia pseudomallei from clinical samples. In acute sepsis, blood is the preferred specimen for culture and therefore should be the target for a rapid diagnostic tool. A lateral flow immunoassay (LFI) for the detection of B. pseudomallei antigen has been developed. This was compared with molecular detection using the targets T3SS1 and IpxO. Forty-five clinical samples of EDTA blood, which were culture-positive, were tested using both modalities. The LFI had a sensitivity of 40 %, whilst molecular detection had a sensitivity of 20 %. The poor performance of molecular detection has been described previously and is largely related to the use of whole-blood specimens collected into blood tubes containing EDTA. Whilst suboptimal, the LFI would be an adjunct in the rapid diagnosis of melioidosis.

The National Strongyloides Working Group in Australia 10 workshops on: commendations and recommendations

[Extract] Strongyloidiasis, caused by the intestinal helminth Strongyloides stercoralis, is commonly found in developing nations in tropical and subtropical regions. Strongyloidiasis was omitted by the World Health Organization (WHO) as one of the Soil Transmitted Helminths in their Neglected Tropical Diseases Roadmap and can therefore be considered one of the most neglected tropical diseases. Despite its reputation as a disease of developing countries, strongyloidiasis remains an important disease in Australia, particularly for Aboriginal and Torres Strait Islanders, and those living in remote communities.

Application of PCR-Based Tools to Explore Strongyloides Infection in People in Parts of Northern Australia

Strongyloidiasis, which is caused by infection with the nematode Strongyloides stercoralis, is endemic to areas of northern Australia. Diagnosis in this region remains difficult due to the distances between endemic communities and diagnostic laboratories, leading to lengthy delays in stool processing for microscopy and culture. PCR represents a viable solution to this difficulty, having potential for high sensitivity detection of S. stercoralis, even in older, unpreserved faecal samples. We prospectively collected 695 faecal specimens that were submitted to The Townsville Hospital Microbiology Laboratory from the North Queensland region for routine parasitological examination, and subjected them to a Strongyloides sp. real-time (q)PCR. Results were confirmed with a novel nested conventional PCR assay targeting the 18S rRNA gene, followed by single-strand conformation polymorphism analysis (SSCP). Of the 695 specimens tested, S. stercoralis was detected in three specimens (0.4%) by classical parasitological methods (direct microscopy and formyl-ether acetate concentration), whereas 42 positives were detected by qPCR (6.0%). Conventional PCR confirmed the real-time PCR results in 24 of the samples (3.5%). Several apparent false-positive results occurred at higher cycle times (Ct) in the qPCR. Use of real-time PCR in these populations is promising for the enhanced detection of disease and to support eradication efforts.

The laboratory diagnosis of Strongyloides stercoralis

Matthew R Watts, Gemma Robertson and Richard S Bradbury Centre for Infectious Diseases and Microbiology, Pathology West – ICMPR, and Marie Bashir Institute, University of Sydney, Westmead Hospital, Westmead, Sydney, NSW, Australia, Tel: +61 2 9845 6255, Email: matthew.watts@health.nsw.gov.au Melbourne Pathology, Collingwood and James Cook University, Tel: +61 3 9287 7700, Email: gemmajrobertson@gmail.com School of Medical and Applied Sciences, Central Queensland University, Rockhampton, Qld, Australia Corresponding author. Email: r.bradbury@cqu.edu.au

The utility of syphilis point of care testing in remote Queensland communities

The rate of infection with syphilis has been steadily climbing over the past ten years Australia-wide. Remote areas of North Queensland have also experienced a dramatic increase in infection, with a concomitant increase in cases of congenital syphilis. Geographically remote locations pose difficulties with respect to specimen transport, processing, and turnaround time. Point-of-care testing (PoCT) offers a practical solution to many of these issues, though it is not a panacea. We aimed to assess the role of syphilis PoCT for remote areas in North Queensland. Throughout 2013–2014 we performed PoCT on field and stored specimens using the SD Bioline Syphilis 3.0 and the Chembio DPP Syphilis Screen and Confirm Assay. Both assays are immunochromatographic card tests, but the Chembio assay detects both non-treponemal and treponemal antibodies. Results were compared to gold standard testing. While both tests are convenient and easy to read, neither is sufficiently sensitive nor specific to replace the gold standard of formal treponemal testing. This is especially true in areas with a low prevalence of disease. Currently our recommendation is that all patients at high risk of infection should be assessed with gold standard testing, even in the presence of a negative point-of-care test.

Enterocytozoon bieneusi genotypes in people with gastrointestinal disorders in Queensland and Western Australia

Enterocytozoon bieneusi is the commonest pathogenic microsporidian found in humans and animals in many countries, but there is scant information on this pathogen in Australia. Here, we conducted the first molecular epidemiological investigation of E. bieneusi in humans with gastrointestinal disorders in Queensland and Western Australia. Genomic DNAs derived from 605 individual faecal samples from children (n = 279) and adults (n = 326) were extracted, and then subjected to nested PCR-based sequencing of the internal transcribed spacer (ITS) of nuclear ribosomal DNA to detect and characterise E. bieneusi. Enterocytozoon bieneusi was detected in eight of 605 human faecal samples (1.3%), including five children (≤3 years of age) and one adult (58 years) in Queensland, and two children (≤3 years) in Western Australia. Analysis of ITS sequence data revealed two known zoonotic (ALP1 and Ind4) and three novel (Hum_q1-3) genotypes of E. bieneusi. Genotype ALP1 identified here in humans has been found previously in farmed alpacas in Australia. Phylogenetic analysis showed that genotypes ALP1, Hum_q1-2 and Ind4 belonged to E. bieneusi Group 1 (with zoonotic potential), whereas genotype Hum_q3 clustered within E. bieneusi Group 10, suggesting that some genotypes within Group 10 might have zoonotic potential. Further investigations of humans, alpacas, marsupials and other animals in Australia will be significant to understand the epidemiology of E. bieneusi in Australia, to identify possible reservoirs of human infection, and to assist in the prevention and control of human microsporidiosis.

Application of real-time PCR for the diagnosis of strongyloidiasis in North Queensland

Aim: Currently most medical schools use an integrated multidisciplinary approach in their curricula; therefore creating a huge challenge to engage medical students by delivering traditional pathology modules. In this study, we aimed to implement multiple advanced strategies to improve medical students’ pathology learning experience at Griffith University. Methods: Students enrolled in the second year of Griffith medical programme between 2011 and 2012 were invited to complete questionnaires rating the value and impact of resources delivered on their learning experience. In total, 272/ 290 students responded. The strategies adopted include virtual microscopy, web-based digitalised interactive modules, clinical scenario-integrated lectures, practical histology sessions and gross specimen demonstration. Quality and usefulness of the delivery of these modules were assessed using a 5 scale questionnaire. Results: In both years, overall score was high (mean score >4.5/ 5) for the histology lectures, clinical integrations and virtual microscopy sessions. The traditional delivery of practical and lecture sessions received lower scores. Qualitative comments suggested that the advanced methods were extremely useful for students’ learning experience of pathology. Discussion: A multidisciplinary approach by clinico-pathological integration and the use of virtual microscopy has the potential to better engage students to pathology learning in the modern medical curriculum.

The use of social media as a 'leadership behaviour' in medicine

School of Medicine and Dentistry, James Cook University, Townsville, Qld 4814, Australia. Email: malcolm.forbes@my.jcu.edu.au; gemmajrobertson@gmail.com Discipline of Psychiatry, University of Adelaide, 55 Frome Road, Adelaide, SA 5005, Australia. The Townsville Hospital, Queensland Health, 100 Angus Smith Drive, Douglas, Qld 4814, Australia. These authors contributed equally to this work. Corresponding author. Email: harris.eyre@gmail.com

Point-of-care testing field trial in mount ISA using the SD bioline syphilis 3.0

Background/Aims: Syphilis is a sexually transmitted illness (STI) caused by Treponema pallidum subsp. pallidum. Effective treatment is achieved with benzathine penicillin, but there are often impediments to follow-up once the laboratory provides a positive diagnosis. Point-of-care testing (PoCT) allows for diagnosis and treatment in a single visit. Methods: A field trial was performed using the SD Bioline Syphilis 3.0 lateral flow assay on 238 patients recruited during the Mount Isa Youth Health Screen. The tests were performed on serum on site and then repeated in The Townsville Hospital microbiology laboratory. The results were compared to enzyme immunoassay (EIA), Treponema pallidum particle agglutination (TPPA) and rapid plasma reagin (RPR) performed on the same samples. Results: Of 238 specimens, 27 were positive by the gold standard of EIA; 22 of the 27 were positive by SD Bioline and no false positives were recorded. The sensitivity of the test is 81.5%, while the specificity is 100%. Discussion: PoCT offers a significant advantage in the management of syphilis. This is especially useful in remote areas where access to health care can be problematic and provision of laboratory results may be delayed. The low sensitivity of this test currently precludes its use as a single diagnostic agent; however, further testing on a larger number of isolates may improve the results.