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Fusarium crown rot (FCR) causes significant grain yield loss in winter cereals around the world. Breeding for resistance and/or tolerance to FCR has been slow with relatively limited success. In this study, multi-species experiments were used to demonstrate an improved method to quantify FCR infection levels at plant maturity using qPCR, as well as the genotype yield retention using residual regression deviation. Using qPCR to measure FCR infection allowed a higher degree of resolution between genotypes than traditional visual stem basal browning assessments. The results were consistent across three environments with different levels of disease expression. The improved measure of FCR infection along with genotype yield retention allows for partitioning of both tolerance and partial resistance. Together these methods offer new insights to FCR partial resistance and its relative importance to tolerance in bread wheat and barley. This new approach offers a more robust, cost-effective way to select for both FCR traits within breeding programs.Key messageGenetic gain for tolerance and partial resistance against Fusarium crown rot (FCR) in winter cereals has been impeded by laborious and variable visual measures of infection severity. This paper presents results of an improved method to quantify FCR infection that are strongly correlated to yield loss and reveal previously unrecognised partial resistance in barley and wheat varieties.

Human activity impacts the evolutionary trajectories of many species worldwide. Global trade of agricultural goods contributes to the dispersal of pathogens reshaping their genetic makeup and providing opportunities for virulence gains. Understanding how pathogens surmount control strategies and cope with new climates is crucial to predicting the future impact of crop pathogens. Here, we address this by assembling a global thousand-genome panel of Zymoseptoria tritici, a major fungal pathogen of wheat reported in all production areas worldwide. We identify the global invasion routes and ongoing genetic exchange of the pathogen among wheat-growing regions. We find that the global expansion was accompanied by increased activity of transposable elements and weakened genomic defenses. Finally, we find significant standing variation for adaptation to new climates encountered during the global spread. Our work shows how large population genomic panels enable deep insights into the evolutionary trajectory of a major crop pathogen.

Zymoseptoria tritici, the causal agent of Septoria tritici blotch, is a fungal wheat pathogen that causes significant global yield losses. Within Z. tritici populations, quantitative differences in virulence among different isolates are commonly observed; however, the genetic components that underpin these differences remain elusive. In this study, intraspecific comparative transcriptomic analysis was used to identify candidate genes that contribute to differences in virulence on the wheat cultivar WW2449. This led to the identification of a multicopy gene that was not expressed in the high‐virulence isolate when compared to the medium‐ and low‐virulence isolates. Further investigation suggested this gene resides in a 7.9‐kb transposon. Subsequent long‐read sequencing of the isolates used in the transcriptomic analysis confirmed that this gene did reside in an active Class II transposon, which is composed of four genes named REP9‐1 to ‐4. Silencing and overexpression of REP9‐1 in two distinct genetic backgrounds demonstrated that its expression alone reduces the number of pycnidia produced by Z. tritici during infection. The REP9‐1 gene identified within a Class II transposon is the first discovery of a gene in a transposable element that influences the virulence of Z. tritici. This discovery adds further complexity to genetic loci that contribute to quantitative virulence in this important pathogen., The expression of the gene REP9‐1, which resides in an active Class II transposon, negatively regulates the rate of pycnidia appearance in planta of the wheat pathogen Zymoseptoria tritici.

Septoria tritici blotch (STB) has been ranked the third most important wheat disease in the world, threatening a large area of wheat production. Although major genes play an important role in the protection against Zymoseptoria tritici infection, the lifespan of their resistance unfortunately is very short in modern agriculture systems. Combinations of quantitative resistance with minor effects, therefore, are believed to have prolonged and more durable resistance to Z. tritici. In this study new quantitative trait loci (QTLs) were identified that are responsible for seedling-stage resistance and adult-plant stage resistance (APR). More importantly was the characterisation of a previously unidentified QTL that can provide resistance during different stages of plant growth or multi-stage resistance (MSR). At the seedling stage, we discovered a new isolate-specific QTL, QSt.wai.1A.1. At the adult-plant stage, the new QTL QStb.wai.6A.2 provided stable and consistent APR in multiple sites and years, while the QTL QStb.wai.7A.2 was highlighted to have MSR. The stacking of multiple favourable MSR alleles was found to improve resistance to Z. tritici by up to 40%.Key messageAn Australian GWAS panel discovered three new QTLs associated with seedling-stage resistance, adult-plant stage resistance, and multi-stage resistance, respectively.

Convergent evolution leads to identical phenotypic traits in different species or populations. Convergence can be driven by standing variation allowing selection to favor identical alleles in parallel or the same mutations can arise independently. However, the molecular basis of such convergent adaptation remains often poorly resolved. Pesticide resistance in agricultural ecosystems is a hallmark of convergence in phenotypic traits. Here, we analyze the major fungal pathogen Zymoseptoria tritici causing serious losses on wheat and with parallel fungicide resistance emergence across continents. We sampled three population pairs each from a different continent spanning periods early and late in the application of fungicides. To identify causal loci for resistance, we combined knowledge from molecular genetics work and performed genome-wide association studies (GWAS) on a global set of isolates. We discovered yet unknown factors in azole resistance including membrane stability functions. We found strong support for the ‘hotspot’ model of resistance evolution with parallel changes in a small set of loci but additional loci showed more population-specific allele frequency changes. Genome-wide scans of selection showed that half of all known resistance loci were overlapping a selective sweep region. Hence, the application of fungicides was one of the major selective agents acting on the pathogen over the past decades. Furthermore, loci identified through GWAS showed the highest overlap with selective sweep regions underlining the importance to map phenotypic trait variation in evolving populations. Our population genomic analyses showed that both de novo mutations and gene flow likely contributed to the parallel emergence of resistance.

Septoria tritici blotch (STB), caused by the fungus Zymoseptoria tritici, is one of the most economically important diseases of wheat. Recently, both factors of a gene‐for‐gene interaction between Z. tritici and wheat, the wheat receptor‐like kinase Stb6 and the Z. tritici secreted effector protein AvrStb6, have been identified. Previous analyses revealed a high diversity of AvrStb6 haplotypes present in earlier Z. tritici isolate collections, with up to c.18% of analysed isolates possessing the avirulence isoform of AvrStb6 identical to that originally identified in the reference isolate IPO323. With Stb6 present in many commercial wheat cultivars globally, we aimed to assess potential changes in AvrStb6 genetic diversity and the incidence of haplotypes allowing evasion of Stb6‐mediated resistance in more recent Z. tritici populations. Here we show, using targeted resequencing of AvrStb6, that this gene is universally present in field isolates sampled from major wheat‐growing regions of the world in 2013–2017. However, in contrast to the data from previous AvrStb6 population studies, we report a complete absence of the originally described avirulence isoform of AvrStb6 amongst modern Z. tritici isolates. Moreover, a remarkably small number of haplotypes, each encoding AvrStb6 protein isoforms conditioning virulence on Stb6‐containing wheat, were found to predominate among modern Z. tritici isolates. A single virulence isoform of AvrStb6 was found to be particularly abundant throughout the global population. These findings indicate that, despite the ability of Z. tritici to sexually reproduce on resistant hosts, AvrStb6 avirulence haplotypes tend to be eliminated in subsequent populations., A snapshot of AvrStb6 isoforms diversity amongst earlier versus more recent natural Zymoseptoria tritici populations revealed a marked decrease in diversity and a global shift to virulence on Stb6 wheat.

Fungal diseases of plants are responsible for major losses in agriculture, highlighting the need for rapid and accurate identification of plant pathogens. Disease outcomes are often defined not only by the main pathogen but are influenced by diverse microbial communities known as the microbiome at sites of infection. Here we present the first use of whole genome shot-gun sequencing with a portable DNA sequencing device as a method for the detection of fungal pathogens from wheat(Triticum aestivum)in a standard molecular biology laboratory. The data revealed that our method is robust and applicable to the diagnosis of fungal diseases including wheat stripe rust (caused byPuccinia striiformisf. sp.tritici),septoria tritici blotch (caused byZymoseptoria tritici)and yellow leaf spot (caused byPyrenophora tritici repentis).We also identified the bacterial genusPseudomonasco-present withPucciniaandZymoseptoriabut notPyrenophorainfections. One limitation of the method is the over-representation of redundant wheat genome sequences from samples. This could be addressed by long-range amplicon-based sequencing approaches in future studies, which specifically target non-host organisms. Our work outlines a new approach for detection of a broad range of plant pathogens and associated microbes using a portable sequencer in a standard laboratory, providing the basis for future development of an on-site disease monitoring system.

Water-soluble carbohydrate accumulation can be selected in wheat breeding programs with consideration of genetic × environmental interactions and relationships with other important characteristics such as relative maturity and nitrogen concentration, although the correlation between WSC traits and grain yield is low and inconsistent. The potential to increase the genetic capacity for water-soluble carbohydrate (WSC) accumulation is an opportunity to improve the drought tolerance capability of rainfed wheat varieties, particularly in environments where terminal drought is a significant constraint to wheat production. A population of elite breeding germplasm was characterized to investigate the potential for selection of improved WSC concentration and total amount in water deficit and well-watered environments. Accumulation of WSC involves complex interactions with other traits and the environment. For both WSC concentration (WSCC) and total WSC per area (WSCA), strong genotype × environment interactions were reflected in the clear grouping of experiments into well-watered and water deficit environment clusters. Genetic correlations between experiments were high within clusters. Heritability for WSCC was larger than for WSCA, and significant associations were observed in both well-watered and water deficit experiment clusters between the WSC traits and nitrogen concentration, tillering, grains per m2, and grain size. However, correlations between grain yield and WSCC or WSCA were weak and variable, suggesting that selection for these traits is not a better strategy for improving yield under drought than direct selection for yield.

Improving water-use efficiency by incorporating drought avoidance traits into new wheat varieties is an important objective for wheat breeding in water-limited environments. This study uses genome wide association studies (GWAS) to identify candidate loci for water-soluble carbohydrate accumulation—an important drought-avoidance characteristic in wheat. Phenotypes from a multi-environment trial with experiments differing in water availability and separate single nucleotide polymorphism (SNP) and diversity arrays technology (DArT) marker sets were used to perform the analyses. Significant associations for water-soluble carbohydrate accumulation were identified on chromosomes 1A, 1B, 1D, 2D, and 4A. Notably, these loci did not collocate with the major loci identified for relative maturity. Loci on chromosome 1D collocated with markers previously associated with the high molecular weight glutenin Glu-D1 locus. Genetic × environmental interactions impacted the results strongly, with significant associations for carbohydrate accumulation identified only in the water-deficit experiments. The markers associated with carbohydrate accumulation may be useful for marker-assisted selection of drought tolerance in wheat.

This work dissects the tripartite horizontal transfer of ToxA, a gene that has a direct negative impact on global wheat yields. Defining the extent of horizontally transferred DNA is important because it can provide clues to the mechanisms that facilitate HGT. Our analysis of ToxA and its surrounding 14 kb suggests that this gene was horizontally transferred in two independent events, with one event likely facilitated by a type II DNA transposon. These horizontal transfer events are now in various processes of decay in each species due to the repeated insertion of new transposons and subsequent rounds of targeted mutation by a fungal genome defense mechanism known as repeat induced point mutation. This work highlights the role that HGT plays in the evolution of host adaptation in eukaryotic pathogens. It also increases the growing body of evidence indicating that transposons facilitate adaptive HGT events between fungi present in similar environments and hosts., Most known examples of horizontal gene transfer (HGT) between eukaryotes are ancient. These events are identified primarily using phylogenetic methods on coding regions alone. Only rarely are there examples of HGT where noncoding DNA is also reported. The gene encoding the wheat virulence protein ToxA and the surrounding 14 kb is one of these rare examples. ToxA has been horizontally transferred between three fungal wheat pathogens (Parastagonospora nodorum, Pyrenophora tritici-repentis, and Bipolaris sorokiniana) as part of a conserved ∼14 kb element which contains coding and noncoding regions. Here we used long-read sequencing to define the extent of HGT between these three fungal species. Construction of near-chromosomal-level assemblies enabled identification of terminal inverted repeats on either end of the 14 kb region, typical of a type II DNA transposon. This is the first description of ToxA with complete transposon features, which we call ToxhAT. In all three species, ToxhAT resides in a large (140-to-250 kb) transposon-rich genomic island which is absent in isolates that do not carry the gene (annotated here as toxa−). We demonstrate that the horizontal transfer of ToxhAT between P. tritici-repentis and P. nodorum occurred as part of a large (∼80 kb) HGT which is now undergoing extensive decay. In B. sorokiniana, in contrast, ToxhAT and its resident genomic island are mobile within the genome. Together, these data provide insight into the noncoding regions that facilitate HGT between eukaryotes and into the genomic processes which mask the extent of HGT between these species.

The disease scald of barley is caused by the pathogen Rhynchosporium commune and can cause up to 30-40% yield loss in susceptible varieties. In this study, the Australian barley cultivar Yerong was demonstrated to have resistance that differed from Turk (Rrs1) based on seedling tests with 11 R. commune isolates. A doubled haploid population with 177 lines derived from a cross between Yerong and Franklin was used to identify quantitative trait loci (QTL) for scald resistance. Scald resistance against four pathogen isolates was assessed at the seedling growth stage in a glasshouse experiment and at the adult growth stage in field experiments with natural infection over three consecutive years. A QTL on chromosome 3H was identified with large effect, consistent with a major gene conferring scald resistance at the seedling stage. Under field conditions, scald percentage was negatively correlated with early relative maturity. A bivariate analysis was used to model scald percentage and relative maturity together, residuals from the regression of scald percentage on relative maturity were used as our phenotype for QTL analysis. This analysis identified one major QTL on chromosome 3H, which mapped to the same position as the QTL identified for scald resistance at seedling stage. The identified QTL on 3H is proposed to be different from the Rrs1 on the basis of seedling resistance against different R. commune isolates and physical map position. The analysis also identified an additional novel QTL on chromosome 7H. This study increases the current understanding of scald resistance and identifies genetic material possessing QTLs useful for the marker-assisted selection of scald resistance in barley breeding programs.

Most known examples of horizontal gene transfer (HGT) between eukaryotes are ancient. These events are identified primarily using phylogenetic methods on coding regions alone. Only rarely are there examples of HGT where non-coding DNA is also reported. The gene encoding the wheat virulence protein ToxA and surrounding 14 kb is one of these rare examples.ToxAhas been horizontally transferred between three fungal wheat pathogens (Parastagonospora nodorum, Pyrenophora tritici-repentisandBipolaris sorokiniana) as part of a conserved ∼14kb element, which contains coding and non-coding regions. Here we use long-read sequencing to define the extent of HGT between these three fungal species. Construction of near-chromosomal level assemblies enabled identification of terminal inverted repeats on either end of the 14kb region, typical of a Type II DNA transposon. This is the first description ofToxAwith complete transposon features, which we call ToxhAT. In all three species, ToxhAT resides in a large (140-250 kb) transposon-rich genomic island which is absent intoxA-isolates. We demonstrate that the horizontal transfer of ToxhAT betweenPyrenophora tritici-repentisandP. nodorumoccurred as part of a large ∼80kb HGT which is now undergoing extensive decay. In contrast, inB. sorokinianaToxhAT and its resident genomic island are mobile within the genome. Together these data provide insight into the non-coding regions that facilitate HGT between eukaryotes and the genomic processes which mask the extent of HGT between these species.IMPORTANCEThis work dissects the tripartite horizontal transferof ToxA; a gene that has a direct negative impact on global wheat yields. Defining the extent of horizontally transferred DNA is important because it can provide clues as to the mechanisms that facilitate HGT. Our analysis ofToxAand its surrounding 14kb suggests that this gene was horizontally transferred in two independent events, with one event likely facilitated by a Type II DNA transposon. These horizontal transfer events are now in various processes of decay in each species due to the repeated insertion of new transposons and subsequent rounds of targeted mutation by a fungal genome defense mechanism known as repeat induced point-mutation. This work highlights the role that HGT plays in the evolution of host adaptation in eukaryotic pathogens. It also increases the growing body of evidence that transposons facilitate adaptive HGT events between fungi present in similar environments and hosts.DATA AVAILABILITYAll raw sequencing data is available under NCBI BioProject PRJNA505097.TheP. nodorumSN15 Whole Genome Shotgun project has been deposited at DDBJ/ENA/GenBank under the accession SSHU00000000. The version SSHU01000000 is described in this paper. TheP. nodorumSN79-1087 Whole Genome Shotgun project has been deposited under the accessionsCP039668-CP039689.The Whole Genome shotgun project and accession numbers forB. sorokinianaisolates are as follows: CS10; SRZH00000000, CS27; SRZG00000000, WAI2406; SRZF00000000, WAI2411; SRZE00000000. Transposon annotations, CS10 and CS27 gene annotations are available athttps://github.com/megancamilla/Transposon-Mediated-transfer-of-ToxA

Zymoseptoria tritici is a globally distributed fungal pathogen which causes Septoria tritici blotch on wheat. Management of the disease is attempted through the deployment of resistant wheat cultivars and the application of fungicides. However, fungicide resistance is commonly observed in Z. tritici populations, and continuous monitoring is required to detect breakdowns in fungicide efficacy. We recently reported azole-resistant isolates in Australia; however, it remained unknown whether resistance was brought into the continent through gene flow or whether resistance emerged independently. To address this question, we screened 43 isolates across five Australian locations for azole sensitivity and performed whole-genome sequencing on 58 isolates from seven locations to determine the genetic basis of resistance. Population genomic analyses showed extremely strong differentiation between the Australian population recovered after azoles began to be used and both Australian populations recovered before azoles began to be used and populations on different continents. The apparent absence of recent gene flow between Australia and other continents suggests that azole fungicide resistance has evolved de novo and subsequently spread within Tasmania. Despite the isolates being distinct at the whole-genome level, we observed combinations of nonsynonymous substitutions at the CYP51 locus identical to those observed elsewhere in the world. We observed nine previously reported nonsynonymous mutations as well as isolates that carried a combination of the previously reported L50S, S188N, A379G, I381V, Y459DEL, G460DEL, and N513K substitutions. Assays for the 50% effective concentration against a subset of isolates exposed to the tebuconazole and epoxiconazole fungicides showed high levels of azole resistance. The rapid, parallel evolution of a complex CYP51 haplotype that matches a dominant European haplotype demonstrates the enormous potential for de novo resistance emergence in pathogenic fungi. IMPORTANCE Fungicides are essential to control diseases in agriculture because many crops are highly susceptible to pathogens. However, many pathogens rapidly evolve resistance to fungicides. A large body of studies have described specific mutations conferring resistance and have often made inferences about the origins of resistance based on sequencing data from the target gene alone. Here, we show the de novo acquisition of resistance to the ubiquitously used azole fungicides in genetically isolated populations of the wheat pathogen Zymoseptoria tritici in Tasmania, Australia. We confirm evidence for parallel evolution through genome-scale analyses of representative worldwide populations. The emergence of complex resistance haplotypes following a well-documented recent introduction of azoles into Australian farming practices demonstrates how rapidly chemical resistance evolves in agricultural ecosystems.

Zymoseptoria tritici is a host-specific, necrotrophic pathogen of wheat. Infection by Z. tritici is characterized by its extended latent period, which typically lasts 2 wks, and is followed by extensive host cell death, and rapid proliferation of fungal biomass. This work characterizes the level of genomic variation in 13 isolates, for which we have measured virulence on 11 wheat cultivars with differential resistance genes. Between the reference isolate, IPO323, and the 13 Australian isolates we identified over 800,000 single nucleotide polymorphisms, of which ∼10% had an effect on the coding regions of the genome. Furthermore, we identified over 1700 probable presence/absence polymorphisms in genes across the Australian isolates using de novo assembly. Finally, we developed a gene tree sorting method that quickly identifies groups of isolates within a single gene alignment whose sequence haplotypes correspond with virulence scores on a single wheat cultivar. Using this method, we have identified

Winter cereal viruses can cause significant crop losses; however, detailed knowledge of their occurrence in New South Wales, Australia is very limited. This paper reports on the occurrence of Wheat streak mosaic virus (WSMV), Wheat mosaic virus (WMoV), Barley yellow dwarf virus (BYDV), Cereal yellow dwarf virus (CYDV), and their serotypes between 2006 and 2014. Detection of WMoV is confirmed in eastern Australia for the first time. The BYDV and CYDV 2014 epidemic is examined in detail using 139 samples of wheat, barley, and oat surveyed from southern New South Wales. The presence of virus was determined using enzyme-linked immunosorbent assays. The results reveal a high frequency of the serotype Barley yellow dwarf virus - MAV as a single infection present in 27% of samples relative to Barley yellow dwarf virus - PAV in 19% and CYDV in 14%. Clear differences emerged in the infection of different winter cereal species by serotypes of BYDV and CYDV. These results are contrasted to other Australian and international studies.

Zymoseptoria tritici is a globally distributed fungal pathogen which causes Septoria tritici blotch on wheat. Management of the disease is attempted through the deployment of resistant wheat cultivars and the application of fungicides. However, fungicide resistance is commonly observed in Z. tritici populations, and continuous monitoring is required to detect breakdowns in fungicide efficacy. We recently reported azole-resistant isolates in Australia; however, it remained unknown whether resistance was brought into the continent through gene flow or whether resistance emerged independently. To address this question, we screened 43 isolates across five Australian locations for azole sensitivity and performed whole-genome sequencing on 58 isolates from seven locations to determine the genetic basis of resistance. Population genomic analyses showed extremely strong differentiation between the Australian population recovered after azoles began to be used and both Australian populations recovered before azoles began to be used and populations on different continents. The apparent absence of recent gene flow between Australia and other continents suggests that azole fungicide resistance has evolved de novo and subsequently spread within Tasmania. Despite the isolates being distinct at the whole-genome level, we observed combinations of nonsynonymous substitutions at the CYP51 locus identical to those observed elsewhere in the world. We observed nine previously reported nonsynonymous mutations as well as isolates that carried a combination of the previously reported L50S, S188N, A379G, I381V, Y459DEL, G460DEL, and N513K substitutions. Assays for the 50% effective concentration against a subset of isolates exposed to the tebuconazole and epoxiconazole fungicides showed high levels of azole resistance. The rapid, parallel evolution of a complex CYP51 haplotype that matches a dominant European haplotype demonstrates the enormous potential for de novo resistance emergence in pathogenic fungi. IMPORTANCE Fungicides are essential to control diseases in agriculture because many crops are highly susceptible to pathogens. However, many pathogens rapidly evolve resistance to fungicides. A large body of studies have described specific mutations conferring resistance and have often made inferences about the origins of resistance based on sequencing data from the target gene alone. Here, we show the de novo acquisition of resistance to the ubiquitously used azole fungicides in genetically isolated populations of the wheat pathogen Zymoseptoria tritici in Tasmania, Australia. We confirm evidence for parallel evolution through genome-scale analyses of representative worldwide populations. The emergence of complex resistance haplotypes following a well-documented recent introduction of azoles into Australian farming practices demonstrates how rapidly chemical resistance evolves in agricultural ecosystems.

Bipolaris sorokiniana is the causal agent of multiple diseases on wheat and barley and is the primary constraint to cereal production throughout South Asia. Despite its significance, the molecular basis of disease is poorly understood. To address this, the genomes of three Australian isolates of B. sorokiniana were sequenced and screened for known pathogenicity genes. Sequence analysis revealed that the isolate BRIP10943 harboured the ToxA gene, which has been associated previously with disease in the wheat pathogens Parastagonospora nodorum and Pyrenophora tritici-repentis. Analysis of the regions flanking ToxA within B. sorokiniana revealed that it was embedded within a 12-kb genomic element nearly identical to the corresponding regions in P. nodorum and P. tritici-repentis. A screen of 35 Australian B. sorokiniana isolates confirmed that ToxA was present in 12 isolates. Sequencing of the ToxA genes within these isolates revealed two haplotypes, which differed by a single non-synonymous nucleotide substitution. Pathogenicity assays showed that a B. sorokiniana isolate harbouring ToxA was more virulent on wheat lines that contained the sensitivity gene when compared with a non-ToxA isolate. This work demonstrates that proteins that confer host-specific virulence can be horizontally acquired across multiple species. This acquisition can dramatically increase the virulence of pathogenic strains on susceptible cultivars, which, in an agricultural setting, can have devastating economic and social impacts.

Septoria tritici blotch (STB) caused by the fungus Mycosphaerella graminicola, is one of the most important foliar diseases of wheat (T. aestivum spp., aestivum L.). Various practices such as crop rotation, application of fungicides, and deployment of genetic resistance have been utilised to control this disease and subsequently reduce yield losses. During the last 20 years, significant progress has been made in understanding host-pathogen interaction, inheritance of STB resistance, localisation of loci controlling STB resistance and identification of molecular markers associated with STB resistance in common wheat. We review the progress made on various aspects of molecular breeding for STB resistance especially on mapping and validation of qualitative and quantitative trait loci in common wheat.

Septoria tritici blotch (STB) caused by Mycosphaerella graminicola (anamorph: Septoria tritici), is one of the most important foliar diseases of wheat. We assessed three doubled-haploid (DH) populations derived from Chara (STB-susceptible)/WW2449 (STB-resistant), Whistler (STB-susceptible)/WW1842 (STB-resistant) and Krichauff (STB susceptible)/WW2451 (STB-resistant) for resistance to a single-pycnidium isolate 79.2.1A of M. graminicola at the seedling stage. STB resistance in each of the three DH populations was conditioned by a single major gene designated as StbWW2449, StbWW1842 and StbWW2451. Linkage analyses and physical mapping indicated that the StbWW loci were located on the short arm of chromosome 1B (IBS). Four simple sequence repeat (SSR) markers linked with STB resistance: Xwmc230, Xbarc119b, Xksum045 and Xbarc008 were located to the distal bin of 1BS.sat1BS-4 (FL: 0.52–1.00) in the 1BS physical map. Xwmc230, Xbarc119b and Xksum045 markers, mapped within 7 cM from StbWW were validated for their linkage and predicted the STB resistance with over 94% accuracy in the 79 advanced breeding lines having WW2449 as one of the parents. The marker interval Xwmc230/Xksum045-Xbarc119b also explained up to 38% of the phenotypic variance at the adult plant stage in all three DH mapping populations. These results have proven that SSR markers are useful in monitoring STB resistance both at seedling and adult plant stages and hence are suitable for routine marker-assisted selection in the wheat breeding programs.

The genus Mycosphaerella Johanson contains many pathogens capable of causing a severe impact on the growth of susceptible eucalypt species. The lack of knowledge about which species are present in Tasmania and their potential risk to the plantation industry prompted this study into the Mycosphaerella species occurring on Eucalyptus globulus and Eucalyptus nitens plantations in Tasmania. A total of 36 plantation and ®ve road verge sites of E. globulus and E. nitens were sampled. Five Mycosphaerella species and three species from associated anamorph genera were isolated and identi®ed in Tasmania; Mycosphaerella nubilosa, Mycosphaerella cryptica, Mycosphaerella tasmaniensis, Mycosphaerella grandis, Mycosphaerella vespa, Coniothyrium ovatum, Sonderhenia eucalypticola and Sonderhenia eucalyptorum. The most frequently isolated species with the highest incidence and severity of infection were M. cryptica and M. nubilosa. These two species appear to have the greatest potential to damage juvenile eucalypt plantations in Tasmania. A link between Mycosphaerella vespa and Coniothyrium ovatum is described for the ®rst time.

The erosion of demethylation inhibitor (DMI) fungicides effectiveness over time in Europe has been attributed to mutation sites in the 14α-demethylase encoded by the nuclear gene CYP51 (Cools and Fraaije 2013). These mutations first appeared in Europe in the 1990s, became widespread over the next 20 years, and were also recently reported in North America (Estep et al. 2015; Lucas et al. 2015). Zymoseptoria tritici has been present and causing disease on wheat (Triticum aestivum) in Australia for many decades and the use of fungicides for its control has become more common over the past 15 years. However, no significant changes in the field performance of fungicides have been noted by growers. To investigate possible undetected changes, we sequenced the CYP51 gene of 18 isolates cultured from wheat leaves collected in a commercial field on 1 July 2012 at Inverleigh, Victoria (–38.086743° S; 143.935783° E), and 3 isolates cultured from wheat leaves collected from trial plots on 22 August 2002 at Wagga Wagga,NSW (–35.044544° S; 147.316318° E). The nucleotide sequence of Z. tritici strain ST1 eburicol 14 alpha-demethylase (CYP51) gene (GenBank Accession No. AY730587.1) was used to design a set of three forward and three reverse sequencing primers across the known mutations. The sequence reads were assembled into contigs and checked for complete sequence. To generate an alignment of mature protein sequences, the mature protein sequence of ST1 (CYP51) gene was downloaded from NCBI to use as a reference. CYP51 nucleotide sequences were translated into amino acid sequences using all six possible frame shifts. These were aligned to the mature protein sequence of ST1 (CYP51) to identify the correct frame-shift sequence, enable the removal of intron sequences, and finally generate an amino acid alignment. Nucleotide and amino acid sequences are deposited in GenBank with Accession Nos. KT201543 to KT201563. Sixteen of the isolates from Victoria were carrying the Y137F mutation, one isolate carried the L50S-Y461S mutation, and one the L50S-S188N-N513K. The three isolates from NSW collected in 2002 contained no mutations compared with the reference accession. Fungicide sensitivities for propiconazole were determined at 50% effective concentrations (EC50) using rates of 100, 30, 10, 3, 1, 0.3, 0.1, 0.03, 0.01, 0.003, and 0.001 mg/liter, and the resistance factor (RF) of each isolate was calculated as fold change in the EC50 compared with that of the wild-type strains as per Cools et al. (2011), four replicates per isolate were performed. Results of the phenotypic assay of a subset of nine isolates (four Y137F mutants, one L50S-Y461S, one L50S-S188N-N513K, and three Wild-type) confirmed the elevated EC50 values for the mutations known to cause reduced sensitivity. The Y137F and L50S-Y461S mutants showed EC50 values of 1.08 and 1.03 mg/liter and RF levels in the range of 3.07 and 2.93. respectively, while the wild-type and L50S-S188N-N513K mutant had EC50 of 0.352 and 0.367 mg/liter, respectively. The EC50 values observed for the isolates appear higher than those found by Cools et al. (2011); however, the RF values are lower for both mutations. Further exploration of CYP51 mutations occurring in Z. tritici within the Australian cropping regions will be necessary to establish how widespread the Y137F and L50S-Y461S mutants are, and if others such as the S524T or the V136A, which are associated with higher levels of resistance to DMIs, are also present.

Aluminium (Al3+) toxicity restricts productivity and profitability of wheat (Triticum aestivum L.) crops grown on acid soils worldwide. Continued gains will be obtained by identifying superior alleles and novel Al3+ resistance loci that can be incorporated into breeding programs. We used association mapping to identify genomic regions associated with Al3+ resistance using 1055 accessions of common wheat from different geographic regions of the world and 178 polymorphic diversity arrays technology (DArT) markers. Bayesian analyses based on genetic distance matrices classified these accessions into 12 subgroups. Genome-wide association analyses detected markers that were significantly associated with Al3+ resistance on chromosomes 1A, 1B, 2A, 2B, 2D, 3A, 3B, 4A, 4B, 4D, 5B, 6A, 6B, 7A, and 7B. Some of these genomic regions correspond to previously identified loci for Al3+ resistance, whereas others appear to be novel. Among the markers targeting TaALMT1 (the major Al3+-resistance gene located on chromosome 4D), those that detected alleles in the promoter explained most of the phenotypic variance for Al3+ resistance, which is consistent with this region controlling the level of TaALMT1 expression. These results demonstrate that genome-wide association mapping cannot only confirm known Al3+-resistance loci, such as those on chromosomes 4D and 4B, but they also highlight the utility of this technique in identifying novel resistance loci.

Research into winter cereal breeding in Australia has focused primarily on studying the effects of rainfed environments. These studies typically show large genotype × environment (GE) interactions, and the complexity of these interactions acts as an impediment to the efficient selection of improved varieties. Wheat has been studied extensively; however, there are no published studies on the GE interactions of triticale in Australia under irrigated production systems. We conducted trials on 101 triticale genotypes at two locations over 4 years under intensive irrigated management practices and measured the yield potential, GE interactions, heritability and estimated genetic gain of yield, lodging resistance and several other traits important for breeding triticale. We found that high yield potential exceeding 10 t ha–1 exists in the Australian germplasm tested and that, in these irrigated trials, genotype accounted for a high proportion of the variability in all measured traits. All genetic parameters such as heritability and estimated genetic gain were high compared with rainfed studies. Breeding of triticale with improved yield and lodging resistance for irrigated environments is achievable and can be pursued with confidence in breeding programs.

To extend the production base of durum wheat in Australia, field trials were conducted on seven registered durum varieties across four seasons and six sites in locations where irrigation was supplied during crop growth. The purpose was to determine if the quality of the grain produced met the requirements for good milling and pasta-making quality and to understand the genotype, environment and their interaction in affecting yield and technological quality of the grain and derived pasta. High grain yields and grain protein were obtained, producing large grain weights, low screenings and low percentage of hard vitreous kernels. Yellow colour of semolina and pasta was reduced marginally but dough and other pasta technological characteristics were similar to typical dryland durum production, with some exceptions. Varieties were identified with potential for production under irrigation.

There is a substantial challenge in identifying appropriate cultivars from databases for introduction into a breeding program. We propose an indirect selection procedure that illustrates how strategically designed multi-environment trials, linked to historical performance databases, can identify germplasm to meet objectives of plant breeding programs. Two strategies for indirect selection of germplasm from the International Wheat and Maize Improvement Center’s (CIMMYT) trial database were developed based on reference and probe genotype sets included in the International Adaptation Trial (IAT). The IAT was designed to improve the understanding of relationships among global spring wheat (Triticum spp.) locations. Grain yield (t/ha) data were collated from 183 IAT trials grown in 40 countries (including Australia) between 2001 and 2004. The reference genotype set strategy used the genetic correlations among locations in the IAT to identify locations similar to a target environment. For a key southern Australian breeding location, Roseworthy, the number of cultivars targeted for selection was reduced to 35% of the original 1252. The Irrigated Winter Cereals Trials (2008–09) aimed to identify high yield potential lines in south-eastern Australian irrigated environments. Thirty-five CIMMYT cultivars identified using the reference genotype selection strategy were grown in this trial series. In all trials, the proportion of CIMMYT cultivars in the top 20% yielding lines exceeded the expected proportion, 0.20. The probe genotype strategy utilised contrasting line yield responses to assess the occurrence of soil-borne stresses such as root lesion nematode (Pratylenchus thorneii) and boron toxicity. For these stresses, the number of targeted cultivars was reduced to 25% and 83% of the original 1252, respectively.

Wheat is one of the most important cereal crops of the world. In order to achieve continued genetic gain in wheat improvement programs, an assessment and utilisation of genetic diversity in a wide range of germplasm are required. The Australian Winter Cereal Collection (AWCC, Tamworth) holds over 33 000 accessions of wheat. In this study, we scanned the genome of 1057 accessions of hexaploid common wheat (Triticum aestivum L.) originating from different geographic regions of the world, with 178 polymorphic DArT™ markers. These accessions comprised modern cultivars (MCs), advanced breeding lines (BLs), and landrace cultivars (LCs). Our results indicate that the LCs had higher polymorphic information content (PIC values) than the MCs and BLs. Cluster and principal coordinate analysis based on genetic distance matrices enabled classification of the 1057 accessions into 12 subgroups. The structure of subgroups appeared to be geographically determined and was generally consistent with pedigrees. Molecular analyses revealed that LCs have unique alleles compared with MCs and BLs, which may be useful for the genetic improvement of wheat.

We studied the genetic structure of Mycosphaerella cryptica following natural infection of Eucalyptus globulus in a genetic trial. Results from this study indicated significant genetic variation within, and between, E. globulus families (three cloned and control-crossed F2 and four open-pollinated families). Single-ascospore isolates were sampled from 72 E. globulus trees with contrasting levels of resistance within the trial, 21 E. nitens trees in an adjacent plantation, and five distant Eucalyptus plantation trees (two E. globulus 100–2400km; two E. nitens 50–2200 km; one E. grandis × E. tereticornis hybrid 1500 km from the trial). Deoxyribonucleic acid from these isolates was scored for the presence/absence of 75 random amplified polymorphic DNA (RAPD) loci. Only 18 RAPD genotypes, which appeared to recombine rarely, were present among the 98 isolates, which indicates that M. cryptica is not strictly heterothallic. Cluster analysis using genetic distance revealed that M cryptica genotypes from the field trial grouped into two clusters that matched differences in isolate culture morphology, indicating that the fungal population comprised two distinctive biotypes. The two biotypes differed markedly in their host interactions. Biotype 2 was found only on E. globulus, whereas biotype 1 infected both E. globulus and E. nitens eucalypt species. Within E. globulus, biotype 2 was almost exclusively collected on resistant trees whereas biotype 1 was found on both resistant and susceptible trees. The hypothesis that there could be specialisation of M cryptica biotypes occurring at two levels, among host species and within host species, is discussed.

Mycosphaerella species are fungal leaf pathogens of Eucalyptus globulus, one of the major plantation tree species in temperate regions of the world. We examined the quantitative genetic variation in susceptibility to infection by Mycosphaerella nubilosa in a genetically diverse population ofE. globulus families growing in a field trial in north-west Tasmania. Disease incidence and severity were assessed on juvenile foliage following a heavy epidemic where mean leaf area damage was 34%. Disease incidence was uniform across the trial. Significant genetic variation for susceptibility was detected with a narrow sense heritability of disease severity being the highest yet reported (h2=0.60) for aMycosphaerella disease of eucalypts. M. nubilosa damage had a significant deleterious effect on tree growth at both the phenotypic and genetic level. We suggest that E. globulus has at least two mechanisms involved in avoiding the deleterious effects of this disease, one is through resistance of the juvenile foliage per se and the other is through the ontogenetic switch to the resistant adult foliage. There is ample opportunity to select genotypes ofE. globulus that are relatively resistant to damage and if these are deployed in areas of high disease risk, significant benefits in plantation productivity could be obtained.

The wheat pathogen Zymoseptoria tritici possesses a large number of accessory chromosomes that may be present or absent in its genome. The genome of the reference isolate IPO323 has been assembled to a very high standard and contains 21 full length chromosome sequences, 8 of which represent accessory chromosomes. The IPO323 reference, when combined with low-cost next-generation sequencing and bioinformatics, can be used as a powerful tool to assess the presence or absence of accessory chromosomes. We present an outline of a range of bioinformatics techniques that can be applied to the analysis of presence–absence variation among accessory chromosomes across 13 novel isolates of Z. tritici.