title: Amplicon ITS2-rDNA targeting Angiostrongylus spp. in canoine CSF
design: DNA islated from CSF of dogs with clinical Canine Neural Angiostrongyliasis from Australia. We adopted an ITS-2 rDNA qPCR diagnostic assay for amplicon metabarcoding and Next-Generation Sequencing (NGS) to allow us to determine Angiostrongylus species. The qPCR primers adapted from Fang et al. (2012) target a ~140bp region of ITS-2 rDNA. This region includes a single nucleotide polymorphism (SNP) (G-to-A) at the 83rd position which discriminates Angiostrongylus cantonensis from Angiostrongylus mackerrasae. Amplicons were prepared either with SensiFast SYBR (BioLine) or Q5 (NEB) and submitted for processing for NGS at the Ramaciotti Centre for Genomics, University of New South Wales, Australia using Illumina MiSeq v2 250 PE.