BACKGROUND: Angiostrongylus cantonensis (rat lungworm) is recognised as the leading cause of human eosinophilic meningitis, a serious condition observed when nematode larvae migrate through the CNS. Canine Neural Angiostrongyliasis (CNA) is the analogous disease in dogs. Both humans and dogs are accidental hosts, and a rapid diagnosis is warranted. A highly sensitive PCR based assay is available but often not readily accessible in many jurisdictions. An alternative DNA amplification assay that would further improve accessibility is needed. This study aimed to assess the diagnostic utility of a newly designed LAMP assay to detect DNA of globally distributed and invasive A. cantonensis and Angiostrongylus mackerrasae, the other neurotropic Angiostrongylus species, which is native to Australia. METHODOLOGY/PRINCIPAL FINDINGS: Cerebrospinal fluid (CSF) from dogs with a presumptive diagnosis of A. cantonensis infection (2020-2022) were received for confirmatory laboratory testing and processed for DNA isolation and ultrasensitive Angiostrongylus qPCR targeting AcanR3390. A newly designed LAMP assay targeting the same gene target was directly compared to the reference ultrasensitive qPCR in a diagnostic laboratory setting to determine the presence of A. cantonensis DNA to diagnose CNA. The LAMP assay (Angie-LAMP) allowed the sensitive detection of A. cantonensis DNA from archived DNA specimens (Kappa = 0.81, 95%CI 0.69-0.92; n = 93) and rapid single-step lysis of archived CSF samples (Kappa = 0.77, 95%CI 0.59-0.94; n = 52). Only A. cantonensis DNA was detected in canine CSF samples, and co-infection with A. mackerrasae using amplicon deep sequencing (ITS-2 rDNA) was not demonstrated. Both SYD.1 and AC13 haplotypes were detected using sequencing of partial cox1. CONCLUSIONS/SIGNIFICANCE: The Angie-LAMP assay is a useful molecular tool for detecting Angiostrongylus DNA in canine CSF and performs comparably to a laboratory Angiostrongylus qPCR. Adaptation of single-step sample lysis improved potential applicability for diagnosis of angiostrongyliasis in a clinical setting for dogs and by extension, to humans.

Centre for Veterinary Education Sydney School of Veterinary Science The University of Sydney Sydney New South Wales Australia

Department of Botany and Zoology Faculty of Science Masaryk University Brno Czech Republic

Department of Ecology and Diseases of Zoo Animals Game Fish and Bees Faculty of Veterinary Hygiene and Ecology University of Veterinary Sciences Brno Brno Czech Republic

Department of Veterinary Sciences and CINeZ FAPPZ Czech University of Life Sciences Prague Prague Czech Republic

Institute of Parasitology Biology Center of Czech Academy of Sciences České Budějovice Czech Republic

Parasitology Laboratory Centre for Infectious Diseases and Microbiology Lab Services Level 3 ICPMR Westmead Hospital Westmead New South Wales Australia

Sydney School of Veterinary Science Faculty of Science The University of Sydney Sydney New South Wales Australia

The University of Sydney Institute for Infectious Diseases Sydney New South Wales Australia

Vetnostics Laverty Pathology North Ryde Laboratory Macquarie Park New South Wales Australia

Westmead Clinical School Faculty of Medicine and Health Sciences The University of Sydney Westmead Hospital Westmead New South Wales Australia

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