Recreating an introductory physiology unit in the Core Concepts form: helping students to think like a physiologist.

This article showcases the redesign of an introductory undergraduate vertebrate physiology unit at Murdoch University (BMS107) to promote student mastery of six Core Concepts of Physiology (Michael J, Cliff W, McFarland J, Modell H, Wright A, SpringerLink. The Core Concepts of Physiology: a New Paradigm for Teaching Physiology, 2017). Concepts were selected for their suitability in an introductory physiology unit and their ability to scaffold advanced physiology learning. Innovative curricular and pedagogical approaches were employed to 1) create a Core Concepts structure, 2) sell the Core Concepts approach to students, 3) foreground Core Concepts in learning materials, 4) actively engage students with Core Concepts, 5) revise, and 6) assess Core Concepts understanding. Median student marks and overall satisfaction with the unit were unaffected by the introduction of a Core Concepts approach. Notably, though, there was a 14% increase in student agreement with the statement "I received feedback that helped me to learn." The challenge of the Core Concepts approach was articulated by students, but these novice learners also recognized Core Concepts as a mechanism to focus their understanding of physiology and promote critical thinking. For teaching staff, a Core Concepts approach was a reinvigorating opportunity to apply their expertise to the teaching of introductory physiology. We propose that a strong Core Concepts emphasis, while challenging, is highly rewarding for staff and provides students with a "disciplinary passport" that better prepares them to progress in diverse courses and professions.NEW & NOTEWORTHY This article presents a "how-to" guide for redesigning an introductory physiology unit to emphasize the Core Concepts of Physiology. Detailed descriptions are provided of innovative, scalable, adjustments to content delivery, assessment, learning objectives, and activities. Staff reflections and student experience suggest a strong Core Concepts emphasis, while challenging, can promote critical thinking and develop an understanding of underlying chemical, physical and biological principles.

Human Pluripotent Stem Cell-Derived Organoids as Models of Liver Disease.

BACKGROUND & AIMS: There are few in vitro models for studying the 3-dimensional interactions among different liver cell types during organogenesis or disease development. We aimed to generate hepatic organoids that comprise different parenchymal liver cell types and have structural features of the liver, using human pluripotent stem cells. METHODS: We cultured H1 human embryonic stem cells (WA-01, passage 27-40) and induced pluripotent stem cells (GM23338) with a series of chemically defined and serum-free media to induce formation of posterior foregut cells, which were differentiated in 3 dimensions into hepatic endoderm spheroids and stepwise into hepatoblast spheroids. Hepatoblast spheroids were reseeded in a high-throughput format and induced to form hepatic organoids; development of functional bile canaliculi was imaged live. Levels of albumin and apolipoprotein B were measured in cell culture supernatants using an enzyme-linked immunosorbent assay. Levels of gamma glutamyl transferase and alkaline phosphatase were measured in cholangiocytes. Organoids were incubated with troglitazone for varying periods and bile transport and accumulation were visualized by live-imaging microscopy. Organoids were incubated with oleic and palmitic acid, and formation of lipid droplets was visualized by staining. We compared gene expression profiles of organoids incubated with free fatty acids or without. We also compared gene expression profiles between liver tissue samples from patients with nonalcoholic steatohepatitis (NASH) versus without. We quantified hepatocyte and cholangiocyte populations in organoids using immunostaining and flow cytometry; cholangiocyte proliferation of cholangiocytes was measured. We compared the bile canaliculi network in the organoids incubated with versus without free fatty acids by live imaging. RESULTS: Cells in organoids differentiated into hepatocytes and cholangiocytes, based on the expression of albumin and cytokeratin 7. Hepatocytes were functional, based on secretion of albumin and apolipoprotein B and cytochrome P450 activity; cholangiocytes were functional, based on gamma glutamyl transferase and alkaline phosphatase activity and proliferative responses to secretin. The organoids organized a functional bile canaliculi system, which was disrupted by cholestasis-inducing drugs such as troglitazone. Organoids incubated with free fatty acids had gene expression signatures similar to those of liver tissues from patients with NASH. Incubation of organoids with free fatty acid-enriched media resulted in structural changes associated with nonalcoholic fatty liver disease, such as decay of bile canaliculi network and ductular reactions. CONCLUSIONS: We developed a hepatic organoid platform with human cells that can be used to model complex liver diseases, including NASH.

Evolution of a wild-plant tobamovirus passaged through an exotic host: Fixation of mutations and increased replication.

Tobamovirus is a group of viruses that have become serious pathogens of crop plants. As part of a study informing risk of wild plant virus spill over to crops, we investigated the capacity of a solanaceous-infecting tobamovirus from an isolated indigenous flora to adapt to new exotic hosts. Yellow tailflower mild mottle virus (YTMMV) (genus Tobamovirus, family Virgaviridae) was isolated from a wild plant of yellow tailflower (Anthocercis littoria, family Solanaceae) and initially passaged through a plant of Nicotiana benthamiana, then one of Nicotiana glutinosa where a single local lesion was used to inoculate a N. benthamiana plant. Sap from this plant was used as starting material for nine serial passages through three plant species. The virus titre was recorded periodically, and 85% of the virus genome was sequenced at each passage for each host. Six polymorphic sites were found in the YTMMV genome across all hosts and passages. At five of these, the alternate alleles became fixed in the viral genome until the end of the experiment. Of these five alleles, one was a non-synonymous mutation (U1499C) that occurred only when the virus replicated in tomato. The mutant isolate harbouring U1499C, designated YTMMV-δ, increased its titre over passages in tomato and outcompeted the wild-type isolate when both were co-inoculated to tomato. That YTMMV-δ had greater reproductive fitness in an exotic host than did the wild type isolate suggests YTMMV evolution is influenced by host changes.

A novel member of the Tombusviridae from a wild legume, Gompholobium preissii.

As part of an investigation into viruses of wild plants in Australia, a contiguous sequence of 3935 nucleotides was obtained after shotgun sequencing of RNA isolated from an asymptomatic wild legume, Gompholobium preissii. Phylogenetic analysis of the sequence revealed that it most closely resembled that of Trailing lespedeza virus 1 (TLV1), a virus isolated from a wild legume in America. The proposed virus, named Gompholobium virus A, and TLV1 are genetically closest to viruses in the genera Alphacarmovirus and Pelarspovirus, family Tombusviridae, but they share features distinguishing them from both groups.

Differential responses to virus challenge of laboratory and wild accessions of australian species of nicotiana, and comparative analysis of RDR1 gene sequences.

Nicotiana benthamiana is a model plant utilised internationally in plant virology because of its apparent hyper-susceptibility to virus infection. Previously, others showed that all laboratory accessions of N. benthamiana have a very narrow genetic basis, probably originating from a single source. It is unknown if responses to virus infection exhibited by the laboratory accession are typical of the species as a whole. To test this, 23 accessions of N. benthamiana were collected from wild populations and challenged with one to four viruses. Additionally, accessions of 21 other Nicotiana species and subspecies from Australia, one from Peru and one from Namibia were tested for susceptibility to the viruses, and for the presence of a mutated RNA-dependent RNA polymerase I allele (Nb-RDR1m) described previously from a laboratory accession of N. benthamiana. All Australian Nicotiana accessions tested were susceptible to virus infections, although there was symptom variability within and between species. The most striking difference was that plants of a laboratory accession of N. benthamiana (RA-4) exhibited hypersensitivity to Yellow tailflower mild mottle tobamovirus infection and died, whereas plants of wild N. benthamiana accessions responded with non-necrotic symptoms. Plants of certain N. occidentalis accessions also exhibited initial hypersensitivity to Yellow tailflower mild mottle virus resembling that of N. benthamiana RA-4 plants, but later recovered. The mutant Nb-RDR1m allele was identified from N. benthamiana RA-4 but not from any of 51 other Nicotiana accessions, including wild accessions of N. benthamiana, demonstrating that the accession of N. benthamiana used widely in laboratories is unusual.

Catharanthus mosaic virus: A potyvirus from a gymnosperm, Welwitschia mirabilis.

A virus from a symptomatic plant of the gymnosperm Welwitschia mirabilis Hook. growing as an ornamental plant in a domestic garden in Western Australia was inoculated to a plant of Nicotiana benthamiana where it established a systemic infection. The complete genome sequence of 9636 nucleotides was determined using high-throughput and Sanger sequencing technologies. The genome sequence shared greatest identity (83% nucleotides and 91% amino acids) with available partial sequences of catharanthus mosaic virus, indicating that the new isolate belonged to that taxon. Analysis of the phylogeny of the complete virus sequence placed it in a monotypic group in the genus Potyvirus. This is the first record of a virus from W. mirabilis, the first complete genome sequence of catharanthus mosaic virus determined, and the first record from Australia. This finding illustrates the risk to natural and managed systems posed by the international trade in live plants and propagules, which enables viruses to establish in new regions and infect new hosts.