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Item Comparative analysis of plastid genomes from allopolyploid Tragopogon miscellus and its diploid parents(John Wiley and Sons Ltd on behalf of German Society for Plant Sciences, Royal Botanical Society of the Netherlands, 2025-09-15) Mukhtar U; Newmarch SC; Winkworth RC; Soltis PS; Soltis DE; Tate JA; Wang X-QTragopogon is a model system for the study of recent, recurrent, and reciprocal allopolyploid formation. Recent research has focused on the fates of nuclear genes duplicated in the allopolyploid T. miscellus relative to the parental diploids, T. dubius and T. pratensis. In contrast, little attention has been given to organellar genomes, which interact with the duplicated nuclear genomes via their gene products. Here we reconstructed plastid genomes (plastomes) for representatives of these three species to investigate their structure and variability among natural and synthetic allopolyploids. Genomic libraries were Illumina-sequenced for several individuals of the allopolyploid T. miscellus and its diploid parents. Whole plastomes were assembled from skimmed data with comparative analyses used to quantify structural and nucleotide variation. Tragopogon plastomes have a typical quadripartite structure and are similar in size to those of other Asteraceae. The 12 plastomes were highly similar, sharing ~99.5%–100% identity. In all but one case, the plastome sequence for each of the polyploids was most similar to that of its expected maternal parent. The exception involved a polyploid that unexpectedly had a T. dubius plastome type, likely as the result of backcrossing with its presumed paternal parent. Such backcrossing events may have contributed to the demise of this polyploid population. Plastome sequences can be used to infer the maternal origins of polyploids as well as investigate ongoing population-level dynamics. More fully assessing plastome variation across the geographic distribution of polyploids and their diploid progenitors may provide additional insights into polyploid formation, population dynamics, and subsequent evolution.Item Development of a rapid loop-mediated isothermal amplification assay for the detection of Dothistroma septosporum(MDPI (Basel, Switzerland), 2021-03-19) Myrholm CL; Tomm BD; Heinzelmann R; Feau N; Hamelin RC; McDougal R; Winkworth RC; Ramsfield TD; Moricca S; Panzavolta TA Loop-Mediated Isothermal Amplification (LAMP) assay was developed for the detection of the pine pathogen Dothistroma septosporum (G. Dorog.) M. Morelet. The specificity of the LAMP assay was tested using a selection of pine needle fungi, including Dothistroma pini Hulbary, and Lecanosticta acicola (Thüm.) Syd.; only D. septosporum DNA was amplified by the test. In terms of sensitivity, the assay was able to detect as little as 1 pg of total D. septosporum DNA. This assay enables DNA extracted from diseased host needles to be rapidly tested for the presence of D. septosporum using relatively simple to operate equipment away from a fully equipped molecular biology laboratory.Item Comparative Analyses of Complete Peronosporaceae (Oomycota) Mitogenome Sequences-Insights into Structural Evolution and Phylogeny.(Oxford University Press on behalf of the Society for Molecular Biology and Evolution, 2022-04-19) Winkworth RC; Neal G; Ogas RA; Nelson BCW; McLenachan PA; Bellgard SE; Lockhart PJMembers of the Peronosporaceae (Oomycota, Chromista), which currently consists of 25 genera and approximately 1,000 recognized species, are responsible for disease on a wide range of plant hosts. Molecular phylogenetic analyses over the last two decades have improved our understanding of evolutionary relationships within Peronosporaceae. To date, 16 numbered and three named clades have been recognized; it is clear from these studies that the current taxonomy does not reflect evolutionary relationships. Whole organelle genome sequences are an increasingly important source of phylogenetic information, and in this study, we present comparative and phylogenetic analyses of mitogenome sequences from 15 of the 19 currently recognized clades of Peronosporaceae, including 44 newly assembled sequences. Our analyses suggest strong conservation of mitogenome size and gene content across Peronosporaceae but, as previous studies have suggested, limited conservation of synteny. Specifically, we identified 28 distinct syntenies amongst the 71 examined isolates. Moreover, 19 of the isolates contained inverted or direct repeats, suggesting repeated sequences may be more common than previously thought. In terms of phylogenetic relationships, our analyses of 34 concatenated mitochondrial gene sequences resulted in a topology that was broadly consistent with previous studies. However, unlike previous studies concatenated mitochondrial sequences provided strong support for higher-level relationships within the family.Item The mitogenome of Phytophthora agathidicida: Evidence for a not so recent arrival of the "kauri killing" Phytophthora in New Zealand(PLOS, 2021-05-21) Winkworth RC; Bellgard SE; McLenachan PA; Lockhart PJ; Blair JEPhytophthora agathidicida is associated with a root rot that threatens the long-term survival of the iconic New Zealand kauri. Although it is widely assumed that this pathogen arrived in New Zealand post-1945, this hypothesis has yet to be formally tested. Here we describe evolutionary analyses aimed at evaluating this and two alternative hypotheses. As a basis for our analyses, we assembled complete mitochondrial genome sequences from 16 accessions representing the geographic range of P. agathidicida as well as those of five other members of Phytophthora clade 5. All 21 mitogenome sequences were very similar, differing little in size with all sharing the same gene content and arrangement. We first examined the temporal origins of genetic diversity using a pair of calibration schemes. Both resulted in similar age estimates; specifically, a mean age of 303.0-304.4 years and 95% HPDs of 206.9-414.6 years for the most recent common ancestor of the included isolates. We then used phylogenetic tree building and network analyses to investigate the geographic distribution of the genetic diversity. Four geographically distinct genetic groups were recognised within P. agathidicida. Taken together the inferred age and geographic distribution of the sampled mitogenome diversity suggests that this pathogen diversified following arrival in New Zealand several hundred to several thousand years ago. This conclusion is consistent with the emergence of kauri dieback disease being a consequence of recent changes in the relationship between the pathogen, host, and environment rather than a post-1945 introduction of the causal pathogen into New Zealand.Item Genomic Profiling of Mycobacterium tuberculosis Strains, Myanmar(Centers for Disease Control and Prevention, 2021-11) Aung HL; Nyunt WW; Fong Y; Biggs PJ; Winkworth RC; Lockhart PJ; Yeo TW; Hill PC; Cook GM; Aung STMultidrug resistance is a major threat to global elimination of tuberculosis (TB). We performed phenotypic drug-susceptibility testing and whole-genome sequencing for 309 isolates from 342 consecutive patients who were given a diagnosis of TB in Yangon, Myanmar, during July 2016‒June 2018. We identified isolates by using the GeneXpert platform to evaluate drug-resistance profiles. A total of 191 (62%) of 309 isolates had rifampin resistance; 168 (88%) of these rifampin-resistant isolates were not genomically related, indicating the repeated emergence of resistance in the population, rather than extensive local transmission. We did not detect resistance mutations to new oral drugs, including bedaquiline and pretomanid. The current GeneXpert MTB/RIF system needs to be modified by using the newly launched Xpert MTB/XDR cartridge or line-probe assay. Introducing new oral drugs to replace those currently used in treatment regimens for multidrug-resistant TB will also be useful for treating TB in Myanmar.