Browsing by Author "Littlejohn M"

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A novel frameshift variant in ALS2 associated with segmental axonopathy in Merino sheep
(BioMed Central Ltd, 2025-12-01) Eager KLM; Jolly RD; Manning L; Willet CE; Snell RG; Lehnert K; Mckean NE; Sneddon NW; O’Rourke BA; Dittmer KE; Tammen I; Littlejohn M
Background: Segmental axonopathy is a recessively inherited neurodegenerative disorder that has affected Merino sheep since the early 1930s. Despite its long-standing recognition, the genetic basis of the condition remained unknown. This study aimed to identify the genetic cause of segmental axonopathy and confirm its pathological features to improve diagnostic accuracy and inform breeding strategies. Results: Whole genome sequencing and genotyping of affected and unaffected Merino sheep identified a novel homozygous frameshift variant in the ALS2 gene that segregated with disease. RNA sequencing of cerebellar peduncle tissue confirmed the nonsense consequence on the ALS2 transcript. Histological analysis highlighted the hallmarks of the disease as large, foamy eosinophilic axonal swellings predominantly in the trigeminal ganglia, with additional degenerative changes in both the brain and spinal cord. These findings support the value of targeted sampling of sensory roots of the trigeminal nerve, spinal cord tracts, and dorsal nerve rootlets to enhance diagnostic accuracy. The same ALS2 variant was found across multiple unrelated flocks in both Australia and New Zealand, indicating a broader presence within the fine-wool Merino sheep population. Conclusions: This study identifies a novel ALS2 frameshift variant associated with segmental axonopathy in Merino sheep and provides both genetic and histological evidence supporting its role in disease pathology. The development of a DNA diagnostic test will enable more informed breeding decisions, reduce the prevalence of this condition, and improve animal welfare and productivity in the Merino industry. Moreover, the findings offer a potential large-animal model for exploring early-onset forms of human motor neuron diseases, including amyotrophic lateral sclerosis, in which ALS2 variants are implicated.
Multiple QTL underlie milk phenotypes at the CSF2RB locus
(2018-09-12) Lopdell TJ; Tiplady K; Couldrey C; Johnson TJJ; Keehan M; Davis SR; Harris B; Spelman RJ; Snell R; Littlejohn M
The genomes of precision edited cloned calves show no evidence for off-target events or increased de novo mutagenesis
(BioMed Central Ltd, 2021-06-17) Jivanji S; Harland C; Cole S; Brophy B; Garrick D; Snell R; Littlejohn M; Laible G
BACKGROUND: Animal health and welfare are at the forefront of public concern and the agricultural sector is responding by prioritising the selection of welfare-relevant traits in their breeding schemes. In some cases, welfare-enhancing traits such as horn-status (i.e., polled) or diluted coat colour, which could enhance heat tolerance, may not segregate in breeds of primary interest, highlighting gene-editing tools such as the CRISPR-Cas9 technology as an approach to rapidly introduce variation into these populations. A major limitation preventing the acceptance of CRISPR-Cas9 mediated gene-editing, however, is the potential for off-target mutagenesis, which has raised concerns about the safety and ultimate applicability of this technology. Here, we present a clone-based study design that has allowed a detailed investigation of off-target and de novo mutagenesis in a cattle line bearing edits in the PMEL gene for diluted coat-colour. RESULTS: No off-target events were detected from high depth whole genome sequencing performed in precursor cell-lines and resultant calves cloned from those edited and non-edited cell lines. Long molecule sequencing at the edited site and plasmid-specific PCRs did not reveal structural variations and/or plasmid integration events in edited samples. Furthermore, an in-depth analysis of de novo mutations across the edited and non-edited cloned calves revealed that the mutation frequency and spectra were unaffected by editing status. Cells in culture, however, appeared to have a distinct mutation signature where de novo mutations were predominantly C > A mutations, and in cloned calves they were predominantly T > G mutations, deviating from the expected excess of C > T mutations. CONCLUSIONS: We found no detectable CRISPR-Cas9 associated off-target mutations in the gene-edited cells or calves derived from the gene-edited cell line. Comparison of de novo mutation in two gene-edited calves and three non-edited control calves did not reveal a higher mutation load in any one group, gene-edited or control, beyond those anticipated from spontaneous mutagenesis. Cell culture and somatic cell nuclear transfer cloning processes contributed the major source of contrast in mutational profile between samples.