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Apicomplexa is a large monophyletic phylum of unicellular, parasitic organisms. Reptiles are hosts to both haemosporidian (Haemosporida) and hemogregarine (Eucoccidiorida) Apicomplexan blood parasites. Within reptiles our understanding of their diversity remains limited, with a paucity of information from Australia, despite a high diversity of squamates (snakes and lizards). We provide a preliminary assessment of haemosporidian and hemogregarine diversity occurring in lizards across northern tropical Australia, building on existing data with results from a microscopy and genetic assessment. We screened total of 233 blood slides using microscopy and detected hemogregarines in 25 geckos, 2 skinks and 1 agamid, while haemosporidians were detected in 13 geckos. DNA sequencing of 28 samples of the hemogregarine 18S rRNA (∼900 bp) nuclear gene revealed five lineages of Australian lizard hemogregarines within heteroxenous adeleids. We sequenced 10 samples of Haemosporida mtDNA (cytb & coI: ∼1313 bp) and phylogenetic analysis with 30 previously published sequences revealed that the Australian Haemosporida grouped within the Haemoproteidae but were not supported as a monophyletic clade. Our results demonstrate that there is significant undocumented evolutionary diversity in Australian lizard haemosporidian and hemogregarine parasites, with preliminary evidence of significantly higher infection rates in geckos.

Item does not contain fulltext Developmental and epileptic encephalopathies (DEEs) are severe neurodevelopmental disorders often beginning in infancy or early childhood that are characterized by intractable seizures, abundant epileptiform activity on EEG, and developmental impairment or regression. CACNA1E is highly expressed in the central nervous system and encodes the alpha1-subunit of the voltage-gated CaV2.3 channel, which conducts high voltage-activated R-type calcium currents that initiate synaptic transmission. Using next-generation sequencing techniques, we identified de novo CACNA1E variants in 30 individuals with DEE, characterized by refractory infantile-onset seizures, severe hypotonia, and profound developmental impairment, often with congenital contractures, macrocephaly, hyperkinetic movement disorders, and early death. Most of the 14, partially recurring, variants cluster within the cytoplasmic ends of all four S6 segments, which form the presumed CaV2.3 channel activation gate. Functional analysis of several S6 variants revealed consistent gain-of-function effects comprising facilitated voltage-dependent activation and slowed inactivation. Another variant located in the domain II S4-S5 linker results in facilitated activation and increased current density. Five participants achieved seizure freedom on the anti-epileptic drug topiramate, which blocks R-type calcium channels. We establish pathogenic variants in CACNA1E as a cause of DEEs and suggest facilitated R-type calcium currents as a disease mechanism for human epilepsy and developmental disorders.

Next-generation sequencing (NGS) approaches are increasingly being used to generate multi-locus data for phylogeographic and evolutionary genetics research. We detail the applicability of a restriction enzyme-mediated genome complexity reduction approach with subsequent NGS (DArTseq) in vertebrate study systems at different evolutionary and geographical scales. We present two case studies using SNP data from the DArTseq molecular marker platform. First, we used DArTseq in a large phylogeographic study of the agamid lizardCtenophorus caudicinctus, including 91 individuals and spanning the geographical range of this species across arid Australia. A low-density DArTseq assay resulted in 28 960 SNPs, with low density referring to a comparably reduced set of identified and sequenced markers as a cost-effective approach. Second, we applied this approach to an evolutionary genetics study of a classic frog hybrid zone (Litoria ewingii–Litoria paraewingi) across 93 individuals, which resulted in 48 117 and 67 060 SNPs for a low- and high-density assay, respectively. We provide a docker-based workflow to facilitate data preparation and analysis, then analyse SNP data using multiple methods including Bayesian model-based clustering and conditional likelihood approaches. Based on comparison of results from the DArTseq platform and traditional molecular approaches, we conclude that DArTseq can be used successfully in vertebrates and will be of particular interest to researchers working at the interface between population genetics and phylogenetics, exploring species boundaries, gene exchange and hybridization.