Comparative Analyses of Complete Peronosporaceae (Oomycota) Mitogenome Sequences-Insights into Structural Evolution and Phylogeny.

Members 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.

Genomic Profiling of Mycobacterium tuberculosis Strains, Myanmar.

Multidrug 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.

The mitogenome of Phytophthora agathidicida: Evidence for a not so recent arrival of the "kauri killing" Phytophthora in New Zealand.

Phytophthora 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.

A LAMP at the end of the tunnel: A rapid, field deployable assay for the kauri dieback pathogen, Phytophthora agathidicida.

The root rot causing oomycete, Phytophthora agathidicida, threatens the long-term survival of the iconic New Zealand kauri. Currently, testing for this pathogen involves an extended soil bioassay that takes 14-20 days and requires specialised staff, consumables, and infrastructure. Here we describe a loop-mediated isothermal amplification (LAMP) assay for the detection of P. agathidicida that targets a portion of the mitochondrial apocytochrome b coding sequence. This assay has high specificity and sensitivity; it did not cross react with a range of other Phytophthora isolates and detected as little as 1 fg of total P. agathidicida DNA or 116 copies of the target locus. Assay performance was further investigated by testing plant tissue baits from flooded soil samples using both the extended soil bioassay and LAMP testing of DNA extracted from baits. In these comparisons, P. agathidicida was detected more frequently using the LAMP test. In addition to greater sensitivity, by removing the need for culturing, the hybrid baiting plus LAMP approach is more cost effective than the extended soil bioassay and, importantly, does not require a centralised laboratory facility with specialised staff, consumables, and equipment. Such testing will allow us to address outstanding questions about P. agathidicida. For example, the hybrid approach could enable monitoring of the pathogen beyond areas with visible disease symptoms, allow direct evaluation of rates and patterns of spread, and allow the effectiveness of disease control to be evaluated. The hybrid LAMP bioassay also has the potential to empower local communities to evaluate the pathogen status of local kauri stands, providing information for disease management and conservation initiatives.

Long-lasting modification of soil fungal diversity associated with the introduction of rabbits to a remote sub-Antarctic archipelago.

During the late nineteenth century, Europeans introduced rabbits to many of the sub-Antarctic islands, environments that prior to this had been devoid of mammalian herbivores. The impacts of rabbits on indigenous ecosystems are well studied; notably, they cause dramatic changes in plant communities and promote soil erosion. However, the responses of fungal communities to such biotic disturbances remain unexplored. We used metabarcoding of soil extracellular DNA to assess the diversity of plant and fungal communities at sites on the sub-Antarctic Kerguelen Islands with contrasting histories of disturbance by rabbits. Our results suggest that on these islands, the simplification of plant communities and increased erosion resulting from the introduction of rabbits have driven compositional changes, including diversity reductions, in indigenous soil fungal communities. Moreover, there is no indication of recovery at sites from which rabbits were removed 20 years ago. These results imply that introduced herbivores have long-lasting and multifaceted effects on fungal biodiversity as well as highlight the low resiliency of sub-Antarctic ecosystems.

Phylogeny and colonization history of Pringlea antiscorbutica (Brassicaceae), an emblematic endemic from the South Indian Ocean Province.

The origins and evolution of sub-Antarctic island floras are not well understood. In particular there is uncertainty about the ages of the contemporary floras and the ultimate origins of the lineages they contain. Pringlea R. Br. (Brassicaceae) is a monotypic genus endemic to four sub-Antarctic island groups in the southern Indian Ocean. Here we used sequences from both the chloroplast and nuclear genomes to examine the phylogenetic position of this enigmatic genus. Our analyses confirm that Pringlea falls within the tribe Thelypodieae and provide a preliminary view of its relationships within the group. Divergence time estimates and ancestral area reconstructions imply Pringlea diverged from a South American ancestor ~5 Myr ago. It remains unclear whether the ancestor of Pringlea dispersed directly to the South Indian Ocean Province (SIOP) or used Antarctica as a stepping-stone; what is clear, however, is that following arrival in the SIOP several additional long-distance dispersal events must be inferred to explain the current distribution of this species. Our analyses also suggest that although Pringlea is likely to have inherited cold tolerance from its closest relatives, the distinctive morphology of this species evolved only after it split from the South American lineage. More generally, our results lend support to the hypothesis that angiosperms persisted on the sub-Antarctic islands throughout the Pliocene and Pleistocene. Taken together with evidence from other sub-Antarctic island plant groups, they suggest the extant flora of sub-Antarctic is likely to have been assembled over a broad time period and from lineages with distinctive biogeographic histories.

New insights into Trimezieae (Iridaceae) phylogeny: what do molecular data tell us?

BACKGROUND AND AIMS: The Neotropical tribe Trimezieae are taxonomically difficult. They are generally characterized by the absence of the features used to delimit their sister group Tigridieae. Delimiting the four genera that make up Trimezieae is also problematic. Previous family-level phylogenetic analyses have not examined the monophyly of the tribe or relationships within it. Reconstructing the phylogeny of Trimezieae will allow us to evaluate the status of the tribe and genera and to examine the suitability of characters traditionally used in their taxonomy. METHODS: Maximum parsimony and Bayesian phylogenetic analyses are presented for 37 species representing all four genera of Trimezieae. Analyses were based on nrITS sequences and a combined plastid dataset. Ancestral character state reconstructions were used to investigate the evolution of ten morphological characters previously considered taxonomically useful. KEY RESULTS: Analyses of nrITS and plastid datasets strongly support the monophyly of Trimezieae and recover four principal clades with varying levels of support; these clades do not correspond to the currently recognized genera. Relationships within the four clades are not consistently resolved, although the conflicting resolutions are not strongly supported in individual analyses. Ancestral character state reconstructions suggest considerable homoplasy, especially in the floral characters used to delimit Pseudotrimezia. CONCLUSIONS: The results strongly support recognition of Trimezieae as a tribe but suggest that both generic- and species-level taxonomy need revision. Further molecular analyses, with increased sampling of taxa and markers, are needed to support any revision. Such analyses will help determine the causes of discordance between the plastid and nuclear data and provide a framework for identifying potential morphological synapomorphies for infra-tribal groups. The results also suggest Trimezieae provide a promising model for evolutionary research.

Detection and diversity of pathogenic Vibrio from Fiji.

Here we investigate the diversity of pathogenic Vibrio species in marine environments close to Suva, Fiji. We use four distinct yet complementary analyses - biochemical testing, phylogenetic analyses, metagenomic analyses and molecular typing - to provide some preliminary insights into the diversity of vibrios in this region. Taken together our analyses confirmed the presence of nine Vibrio species, including three of the most important disease-causing vibrios (i.e. V. cholerae, V. parahaemolyticus and V. vulnificus), in Fijian marine environments. Furthermore, since toxigenic V. parahaemolyticus are present on fish for consumption we suggest these bacteria represent a potential public health risk. Our results from Illumina short read sequencing are encouraging in the context of microbial profiling and biomonitoring. They suggest this approach may offer an efficient and cost-effective method for studying the dynamics of microbial diversity in marine environments over time.

Mitochondrial sequence data and Dipsacales phylogeny: mixed models, partitioned Bayesian analyses, and model selection.

Phylogenetic analyses of chloroplast DNA sequences, morphology, and combined data have provided consistent support for many of the major branches within the angiosperm clade Dipsacales. Here we use sequences from three mitochondrial loci to test the existing broad scale phylogeny and in an attempt to resolve several relationships that have remained uncertain. Parsimony, maximum likelihood, and Bayesian analyses of a combined mitochondrial data set recover trees broadly consistent with previous studies, although resolution and support are lower than in the largest chloroplast analyses. Combining chloroplast and mitochondrial data results in a generally well-resolved and very strongly supported topology but the previously recognized problem areas remain. To investigate why these relationships have been difficult to resolve we conducted a series of experiments using different data partitions and heterogeneous substitution models. Usually more complex modeling schemes are favored regardless of the partitions recognized but model choice had little effect on topology or support values. In contrast there are consistent but weakly supported differences in the topologies recovered from coding and non-coding matrices. These conflicts directly correspond to relationships that were poorly resolved in analyses of the full combined chloroplast-mitochondrial data set. We suggest incongruent signal has contributed to our inability to confidently resolve these problem areas.

Viburnum phylogeny based on combined molecular data: implications for taxonomy and biogeography.

We investigated Viburnum phylogeny using separate and combined analyses of DNA sequence data from two chloroplast and three nuclear loci. Separate analyses of nuclear and chloroplast data sets resulted in gene trees that were generally congruent with one another and with trees from two previous analyses. Our gene trees do differ in the position of section Pseudotinus, as well as in species relationships within sections Pseudotinus and Lentago. However, tests for incongruence indicate that differences between the nuclear and chloroplast data are not significant. Furthermore, gene trees from combined analyses were highly similar to those found in separate analyses, suggesting that these localized differences do not affect other parts of the tree. Our analyses provide convincing support for numerous relationships, although there is still uncertainty at the base of the tree. To facilitate future study, we propose informal names for 12 well-supported species groups, as well as for several higher-level clades. We also discuss the biogeographic implications of our phylogeny, focusing on repeated, although apparently temporally incongruent, patterns of disjunction between the Old and New Worlds.

Viburnum phylogeny: evidence from the duplicated nuclear gene GBSSI.

DNA sequencing studies of the granule-bound starch synthase gene (GBSSI) indicate the presence of two loci in Viburnum. Gene trees from separate and combined phylogenetic analyses of the GBSSI paralogues are generally congruent with each other and with trees from previous analyses, especially those of Donoghue et al. [Syst. Bot. 29 (2004) 188] based on nuclear ribosomal ITS and chloroplast trnK intron DNA sequences. Specifically, our GBSSI trees confirm (i) the monophyly of some and non-monophyly of other traditionally recognized taxonomic sections, (ii) the presence of three major supra-sectional lineages within Viburnum, and (iii) the resolution of many species relationships within the section-level clades. Analyses of GBSSI also provide greater resolution of relationships within the largest supra-sectional lineage. Relationships at the base of the Viburnum phylogeny remain uncertain; in particular, the position of the root, relationships among the supra-sectional clades, and the exact placement of several smaller groups (e.g., Viburnum clemensiae, Viburnum urceolatum, and section Pseudotinus). In two lineages each GBSSI paralogue is represented by two distinct sequences. The presence of additional copies appears to be correlated with polyploidy in these clades. Placement of the homoeologues in our gene trees suggests the possibility of a hybrid origin for these polyploids.

The origins and evolution of the genus Myosotis L. (Boraginaceae).

Although morphologically well defined, the phylogeny and taxonomy of Myosotis has been uncertain. In particular it has been unclear whether the genus had a Northern Hemisphere or Australasian origin. However, separate analyses of the ITS and the 3' region of matK, as well as a combined analysis of ITS, 3' matK, the psbA-trnA spacer, and 3' ndhF regions indicate that several distinct lineages exist within Myosotis and strongly support a Northern Hemisphere origin for the genus. Further, the observed transoceanic distributions and levels of genetic divergence between lineages indicate that long distance dispersal has been important for establishing the current geographic range expansion of Myosotis. Our molecular data also suggest that the diversification of Australasian Myosotis has occurred since the late Tertiary and is largely due to radiation within and from New Zealand. This inference is consistent with the findings of recent phylogenetic studies on other New Zealand alpine genera. Our results highlight the important role played by late Tertiary and Quaternary climate change in explaining current floristic diversity. The genetic relationships reported here also suggest that the current infrageneric taxonomy of Myosotis does not fully reflect the evolution of the genus.