Genome-scale phylogenies reveal relationships among Parastagonospora species infecting domesticated and wild grasses.

Several plant pathogenic Parastagonospora species have been identified infecting wheat and other cereals over the past 50 years. As new lineages were discovered, naming conventions grew unwieldy and the relationships with previously recognized species remained unclear. We used genome sequencing to clarify relationships among these species and provided new names for most of these species. Six of the nine described Parastagonospora species were recovered from wheat, with five of these species coming from Iran. Genome sequences revealed that three strains thought to be hybrids between P. nodorum and P. pseudonodorum were not actually hybrids, but rather represented rare gene introgressions between those species. Our data are consistent with the hypothesis that P. nodorum originated as a pathogen of wild grasses in the Fertile Crescent, then emerged as a wheat pathogen via host-tracking during the domestication of wheat in the same region. The discovery of a diverse array of Parastagonospora species infecting wheat in Iran suggests that new wheat pathogens could emerge from this region in the future. Citation: Croll D, Crous PW, Pereira D, et al. 2021. Genome-scale phylogenies reveal relationships among Parastagonospora species infecting domesticated and wild grasses. Persoonia 46: 116-128. https://doi.org/10.3767/persoonia.2021.46.04.

Pyricularia graminis-tritici, a new Pyricularia species causing wheat blast.

Pyricularia oryzae is a species complex that causes blast disease on more than 50 species of poaceous plants. Pyricularia oryzae has a worldwide distribution as a rice pathogen and in the last 30 years emerged as an important wheat pathogen in southern Brazil. We conducted phylogenetic analyses using 10 housekeeping loci for 128 isolates of P. oryzae sampled from sympatric populations of wheat, rice, and grasses growing in or near wheat fields. Phylogenetic analyses grouped the isolates into three major clades. Clade 1 comprised isolates associated only with rice and corresponds to the previously described rice blast pathogen P. oryzae pathotype Oryza (PoO). Clade 2 comprised isolates associated almost exclusively with wheat and corresponds to the previously described wheat blast pathogen P. oryzae pathotype Triticum (PoT). Clade 3 contained isolates obtained from wheat as well as other Poaceae hosts. We found that Clade 3 is distinct from P. oryzae and represents a new species, Pyricularia graminis-tritici (Pgt). No morphological differences were observed among these species, but a distinctive pathogenicity spectrum was observed. Pgt and PoT were pathogenic and highly aggressive on Triticum aestivum (wheat), Hordeum vulgare (barley), Urochloa brizantha (signal grass), and Avena sativa (oats). PoO was highly virulent on the original rice host (Oryza sativa), and also on wheat, barley, and oats, but not on signal grass. We conclude that blast disease on wheat and its associated Poaceae hosts in Brazil is caused by multiple Pyricularia species. Pyricularia graminis-tritici was recently found causing wheat blast in Bangladesh. This indicates that P. graminis-tritici represents a serious threat to wheat cultivation globally.

QoI Resistance and Mitochondrial Genetic Structure of Zymoseptoria tritici in Morocco.

In total, 230 single-conidial isolates of the fungal wheat pathogen Zymoseptoria tritici (formerly Septoria tritici, teleomorph: Mycosphaerella graminicola) were sampled in Morocco in 2008 and 2010 to assess resistance against quinone outside inhibitors (QoIs), a widely used group of fungicides in wheat pest management. All 134 isolates sampled in 2008 were QoI sensitive. In contrast, 9 of the 96 isolates from the 2010 collection were resistant, suggesting a recent emergence of the resistance. Mitochondrial (mt)DNA-sequence analyses identified four haplotypes among the resistant isolates. Wright's F statistics (FST) analyses from mtDNA sequences revealed a shallow population structure of Z. tritici within Morocco and a substantial asymmetric gene flow from Europe into Morocco. A phylogenetic reconstruction including Moroccan and European isolates clustered the haplotypes regardless of their geographic origin. The four Moroccan QoI-resistant mitochondrial haplotypes clustered in two distinct clades in the tree topology, suggesting at least two independent origins of the resistance. This study reported, for the first time, the occurrence of QoI-resistant genotypes of Z. tritici in Morocco. Our findings are consistent with the hypothesis that QoI resistance emerged very recently through parallel genetic adaptation in Morocco, although gene flow from Europe cannot be excluded.

Migration patterns and changes in population biology associated with the worldwide spread of the oilseed rape pathogen Leptosphaeria maculans.

Pathogen introductions into novel areas can lead to the emergence of new fungal diseases of plants. Understanding the origin, introduction pathways, possible changes in reproductive system and population size of fungal pathogens is essential in devising an integrated strategy for the control of these diseases. We used minisatellite markers to infer the worldwide invasion history of the fungal plant pathogen Leptosphaeria maculans, which causes stem canker (blackleg) of oilseed and vegetable brassicas. Clustering analyses partitioned genotypes into distinct populations corresponding to major geographic regions, along with two differentiated populations in Western Canada. Comparison of invasion scenarios using Approximate Bayesian Computation suggested an origin of the pathogen in the USA, the region where epidemics were first recorded, and independent introductions from there over the last few decades into Eastern Canada (Ontario), Europe and Australia. The population in Western Canada appeared to be founded from a source in Ontario and the population in Chile resulted from an admixture between multiple sources. A bottleneck was inferred for the introduction into Western Canada but not into Europe, Ontario or Australia. Clonality appeared high in Western Canada, possibly because environmental conditions there were less conducive to sexual reproduction. Leptosphaeria maculans is a model invasive pathogen with contrasting features in different regions: shallow population structure, high genetic variability and regular sexual recombination in some regions, by comparison with reduced genetic variability, high rates of asexual multiplication, strong population structure or admixture in others.

Habitat-specific population structure in native western flower thrips Frankliniella occidentalis (Insecta, Thysanoptera).

Invasions by pest organisms are among the main challenges for sustainable crop protection. They pose a serious threat to crop production by introducing a highly unpredictable element to existing crop protection strategies. The western flower thrips Frankliniella occidentalis (Insecta, Thysanoptera) managed to invade ornamental greenhouses worldwide within < 25 years. To shed light on possible genetic and/or ecological factors that may have been responsible for this invasion success, we studied the population genetic structure of western flower thrips in its native range in western North America. Analysis of nucleotide sequence variation and variation at microsatellite loci revealed the existence of two habitat-specific phylogenetic lineages (ecotypes) with allopatric distribution. One lineage is associated with hot/dry climates, the second lineage is restricted to cool/moist climates. We speculate that the ecological niche segregation found in this study may be among the key factors determining the invasion potential of western flower thrips.

Frequency of sexual recombination by Mycosphaerella graminicola in mild and severe epidemics.

The importance of sexual recombination in determining fungal population structure cannot be inferred solely from the relative abundance of sexual and asexual spores and reproductive structures. To complement a previously reported study of proportions of Mycosphaerella graminicola ascocarps and pycnidia, we investigated the share of sexual recombinants among isolates randomly derived from the same field at the same time. Early in three successive growing seasons (those ending in 1998, 1999, and 2000), field plots of the susceptible winter wheat cultivar Stephens were inoculated with suspensions of two M. graminicola isolates that each had rare alleles at restriction fragment length polymorphism (RFLP) loci. Near harvest time, leaves were randomly sampled from the same plots, and a population of over 100 monopycnidial isolates was created for each year of the experiment. Natural populations were also sampled from noninoculated plots in the 1999 and 2000 seasons, in order to compare allele frequencies. Based on RFLP haplotypes and DNA fingerprints, isolates from the inoculated plots were categorized by both inspection and Bayesian methods as inoculant clones, recombinants, or immigrants. Inoculation in the 2000 season was delayed, and the recovery rate of inoculant types was just 1 to 2%. In 1998, a high-disease year, and 1999, a low-disease year, inoculants comprised 36 and 22 to 23% of end-of-season samples, respectively. In those 2 years, recombinants as a percentage of inoculant descendants (both sexual and asexual) were 35 and 32%, respectively. By comparison, the study of fruiting bodies had found 93 and 32% of M. graminicola fruiting bodies were ascocarps in 1998 and 1999, respectively. These findings support the hypothesis that sexual recombination makes a relatively consistent contribution to M. graminicola population structure, despite differences in epidemic severity and ascocarp proportions.

Holarctic phylogeography of Arctic charr (Salvelinus alpinus L.) inferred from mitochondrial DNA sequences.

This study evaluated mitochondrial DNA (mtDNA) sequence variation in a 552-bp fragment of the control region of Arctic charr (Salvelinus alpinus) by analyzing 159 individuals from 83 populations throughout the entire range of the complex. A total of 89 (16.1%) nucleotide positions were polymorphic, and these defined 63 haplotypes. Phylogenetic analyses supported the monophyly of the complex and assigned the observed haplotypes to five geographic regions that may be associated with different glacial refugia. Most notably, a formerly defined major evolutionary lineage (S. a. erythrinus) ranging from North America across the Arctic archipelago to the Eurasian continent has now been partitioned into the Arctic group and the newly identified Siberian group. The Beringian group, formed entirely by specimens assigned to S. malma (Dolly Varden), encompassed the area formerly assigned to S. a. taranetzi. The latter, due to a unique haplotype, became the basal member of the Arctic group. Overall, the S. alpinus complex reflects divergent evolutionary groups coupled with shallow intergroup differentiation, also indicated by an analysis of molecular variance that attributed 73.7% (P < 0.001) of the total genetic variance among groups. Time estimates, based on sequence divergence, suggest a separation of the major phylogeographic groups during early to mid-Pleistocene. In contrast, colonization of most of today's range started relatively recently, most likely late Pleistocene during the last retreat of ice sheets some 10,000-20,000 years ago. This time scale obviously is too shallow for detecting significant variation on a smaller scale using mtDNA markers. However, other studies using nuclear microsatellite DNA variation strongly suggested ongoing evolution within groups by revealing strong population-genetic substructuring and restricted gene flow among populations. Thus, Arctic charr could serve as a model organism to investigate the linkage between historical and contemporary components of phylogeographic structuring in fish, and, with a global perspective of the distribution of genetic variation as a framework, meaningful comparisons of charr studies at a smaller geographic scale will now be possible.

Evolution of the CYP51 gene in Mycosphaerella graminicola: evidence for intragenic recombination and selective replacement.

Recent findings are consistent with a slow but constant shift towards reduced sensitivity of Mycosphaerella graminicola to azole fungicides, which target the CYP51 gene. The goal of this study was to elucidate the evolutionary mechanisms through which CYP51-based mutations associated with altered sensitivity have evolved in M. graminicola over space and time. To accomplish this, we sequenced and compared a portion of the CYP51 gene encompassing the main mutations associated with altered sensitivity towards demethylation inhibitor fungicides. The CYP51 gene showed an extraordinary dynamic shift consistent with a selective haplotype replacement both in space and in time. No mutations associated with increased resistance to azoles were found in non-European populations. These mutations were also absent in the oldest collections from Europe, whereas they dominated in the recent European populations. Intragenic recombination was identified as an important evolutionary process in populations affected by high fungicide selection, suggesting the creation of novel alleles among existing mutations as a potential source of novel resistance alleles. We propose that CYP51 mutations giving resistance in M. graminicola arose only locally (perhaps in Denmark or the UK) and were then spread eastward across Europe through wind-dispersed ascospores. We conclude that recurring cycles of recombination coupled with selection due to the widespread use of azole fungicides will increase the frequency of novel mutants or recombinants with higher resistance. Long-distance gene flow due to wind dispersal of ascospores will move the resulting new alleles to new areas following the prevailing wind directions. A selective replacement favouring haplotypes with various coding mutations at the target site for azole fungicides during the last 5-10 years is the most likely cause of the decrease in sensitivity reported for many azole fungicides in the same period.

The origin and colonization history of the barley scald pathogen Rhynchosporium secalis.

The origins of pathogens and their past and present migration patterns are often unknown. We used phylogenetic haplotype clustering in conjunction with model-based coalescent approaches to reconstruct the genetic history of the barley leaf pathogen Rhynchosporium secalis using the avirulence gene NIP1 and its flanking regions. Our results falsify the hypothesis that R. secalis emerged in association with its host during the domestication of barley 10,000 to 15,000 years ago in the Fertile Crescent and was introduced into Europe through the migration of Neolithic farmers. Estimates of time since most recent common ancestor (2500-5000 BP) placed the emergence of R. secalis clearly after the domestication of barley. We propose that modern populations of R. secalis originated in northern Europe following a host switch, most probably from a wild grass onto cultivated barley shortly after barley was introduced into northern Europe. R. secalis subsequently spread southwards into already established European barley-growing areas.

Invasion success of the bumblebee, Bombus terrestris, despite a drastic genetic bottleneck.

In early 1992, the European bumblebee, Bombus terrestris, was first seen in Tasmania and currently has spread to most of the island. Here, we report on the genetic structure, using micro-satellites, of the invading population from samples collected in the years 1998-2000, a few years after the first sighting of the species in its new area. The data show that the Tasmanian population has a very low genetic diversity, with less than half of the allelic richness (Richness=2.89 alleles; H(exp)=0.591) and lower levels of heterozygosity as compared to populations in New Zealand (4.24 alleles; H(exp)=0.729) and Europe (5.08 alleles; H(exp)=0.826). In addition, the genetic data suggest that the invasion must have happened once, probably around late 1991, and was the result of very few, perhaps only two, individuals arriving in Tasmania. Furthermore, these founders came from the New Zealand population. Today, the population in the south of Tasmania seems to act as a source population from which individuals migrate into other parts of the state. A similar source-sink structure seems also the case for New Zealand. The data show that B. terrestris is a highly invasive species capable of establishing itself even after a dramatic genetic bottleneck. B. terrestris may be an invasive species due to the haplo-diploid sex determination system, which exposes recessive, deleterious mutations to selection. Offspring of such purged lines may then be able to tolerate high levels of inbreeding.

Host-associated genetic differentiation in Thrips tabaci (Insecta; Thysanoptera), as determined from mtDNA sequence data.

We tested for host-associated genetic differentiation in 22 populations of Thrips tabaci collected from tobacco and leek, respectively. Clustering analyses and haplotype networks based on sequence variation at a fragment of the mitochondrial cytochrome oxidase I gene yielded three major evolutionary lineages; two were clearly associated with leek and the third with tobacco. These genetic findings corroborated recent experimental observations on the heterogeneity of T. tabaci populations with regard to host-plant preference and their capacity to be vectors for tomato spotted wilt virus. Estimated divergence times suggested an ancient divergence of these lineages dating back to the Miocene 28-21 million years ago. F(ST) values between these lineages ranged between 0.824 and 0.954 (P<0.001 for all comparisons), and sequence divergences ranged between 4 and 11%. Given these findings and by the standards of genetic and ecological differentiation in other published species groups, T. tabaci must be considered a complex of cryptic (sub)species.