Species composition,toxigenic potential and pathogenicity of Fusarium graminearum species complex isolates from southern Brazilian rice

This study aimed to assess the extent and distribution of Fusarium graminearum species complex (FGSC) diversity in rice seeds produced in southern Brazil. Four species and two trichothecene genotypes were detected among 89 FGSC isolates, based on a multilocus genotyping assay: F. asiaticum (69·6%) with the nivalenol (NIV) genotype, F. graminearum (14·6%) with the 15‐acetyldeoxynivalenol (ADON) genotype, and F. cortaderiae (14·6%) and F. meridionale (1·1%), both with the NIV genotype. Seven selected F. asiaticum isolates from rice produced NIV in rice‐based substrate in vitro, at levels ranging from 4·7 to 84·1 μg g^?1. Similarly, two F. graminearum isolates from rice produced mainly 15‐ADON (c. 15–41 μg g^?1) and a smaller amount of 3‐ADON (c. 6–12 μg g^?1). One F. meridionale and two F. cortaderiae isolates did not produce detectable levels of trichothecenes. Two F. asiaticum isolates from rice and two from wheat (from a previous study), and one F. graminearum isolate from wheat, were pathogenic to both crops at various levels of aggressiveness based on measures of disease severity in wheat spikes and rice kernel infection in a greenhouse assay. Fusarium asiaticum and the reference F. graminearum isolate from wheat produced NIV, and deoxynivalenol and acetylates, respectively, in the kernels of inoculated wheat heads. No trichothecene was produced in kernels from inoculated rice panicles by any of the isolates. These findings constitute the first report of FGSC composition in rice outside Asia, and confirm the dominance of F. asiaticum in rice agroecosystems.     

Low genetic diversity of Rhynchosporium commune in Iran,a secondary centre of barley origin

Rhynchosporium commune is a destructive pathogen of barley, causing leaf scald. Previous microsatellite studies used Syria as a representative of cultivated barley's centre of origin, the Fertile Crescent. These suggested that R. commune and Hordeum vulgare (cultivated barley) did not co‐evolve in the host's centre of origin. The present study compares R. commune populations from Syria with those from Iran, which represents a secondary centre of origin for barley at the eastern edge of the Iranian Plateau. Results from this study also suggest that R. commune and barley did not co‐evolve in the centre of origin of cultivated barley. This was evidenced by the low pathogen genetic diversity in Iran, which was even lower than in Syria, indicating that the pathogen may have been introduced recently into Iran, perhaps through infected barley seed. Hierarchical analyses of molecular variance revealed that most genetic diversity in Iran and Syria is distributed within populations, with only 14% among populations. Analyses of multilocus association, genotype diversity and mating type frequency suggest that Iranian populations reproduce predominantly asexually. The presence of both mating types on barley and uncultivated grasses suggest a potential for sexual reproduction. Rhynchosporium commune was also found on Hordeum murinum subsp. glaucum, H. vulgare subsp. spontaneum, Lolium multiflorum and, for the first time, on Avena sativa. The variety of wild grasses that can be infected with R. commune in Iran raises concerns of these grasses acting as evolutionary breeding grounds and sources of inoculum.     

Composition and toxigenic potential of the Fusarium graminearum species complex from maize ears,stalks and stubble in Brazil

A large collection (n = 539) of Fusarium graminearum species complex (FGSC) isolates was obtained from Brazilian maize, and collections formed according to geography and maize part: (i) kernel (n = 110) from south and south‐central Brazil; (ii) stalk (n = 134) from Paraná state (south); and (iii) stubble (n = 295) from Rio Grande do Sul state (south). Species composition, identified using a multilocus genotype approach, was assessed separately in each collection due to differences in geographic sampling. Overall, three species were found: F. meridionale (Fmer; 67% prevalence) with the nivalenol (NIV) genotype, F. graminearum (Fgra; 19%) with the 15‐acetyl (A) deoxynivalenol (DON) genotype, and F. cortaderiae (Fcor; 14%) with the NIV (49/74) or the 3‐ADON (25/74) genotype. In kernels, Fmer was spread across all locations and Fgra and Fcor were found mostly at high elevation (>800 m a.s.l.). The majority (97·8%) of stalk isolates was assigned to Fmer; three were assigned to Fgra. In the stubble, Fmer was less dominant (53%), with a shift towards Fcor as the most frequent species at high elevation sites (>600 m a.s.l.). No differences in the mycelial growth rate were observed among isolates from each species grown at 15°C. Fgra grew faster at 25°C and Fmer showed the widest range of variation across the isolates at both temperatures. The survey data suggest that Fmer may outcompete other species on ears and stalks in comparison to stubble. Additional sampling that controls for other factors, as well as direct testing of aggressiveness on ears and stalk tissue, will be needed to fully evaluate this hypothesis.     

Diversity and pathogenicity of Fusarium graminearum species complex from maize stalk and ear rot strains in northeast China

The Fusarium graminearum species complex (FGSC) is an important group of pathogens distributed in maize‐producing areas worldwide. This study investigated the genetic diversity and pathogenicity of 40 FGSC isolates obtained from stalk rot and ear rot samples collected from 42 locations in northeastern China during 2013 and 2014. A phylogenetic tree of translation elongation factor (EF‐la) sequences designated the 40 isolates as F. graminearum sensu stricto (67.5%) and F. boothii (32.5%). By using inter‐simple sequence repeat analysis (ISSR), it was shown that the isolates were divided into two clades, which corresponded to the species identity of the isolates. However, the isolates from the two different diseases could not be distinguished in pathogenicity. The disease severity index of seedlings inoculated with stalk isolates was slightly higher than that of seedlings inoculated with isolates from infected ears, whereas the pathogenicity of the stalk and ear isolates were identical.     

Pathogenicity of Fusarium graminearum and F. meridionale on soybean pod blight and trichothecene accumulation

Soybean (Glycine max) is the most important crop in Argentina. At present Fusarium graminearum is recognized as a primary pathogen of soybean in several countries in the Americas, mainly causing seed and root rot and pre‐ and post‐emergence damping off. However, no information about infections at later growth stages of soybean development and pathogenicity of F. graminearum species complex is available. Therefore, the objectives of this study were to compare the pathogenicity of F. graminearum and F. meridionale isolates towards soybean under field conditions and to evaluate the degree of pathogenicity and trichothecene production of these two phylogenetic species that express different chemotypes. Six isolates of F. graminearum and F. meridionale were evaluated during 2012/13 and 2013/14 soybean growing seasons for pod blight severity, percentage of seed infected in pods and kernel weight reduction. The results showed a higher aggressiveness of both F. graminearum and F. meridionale species during the 2013/14 season. However, the differences in pathogenicity observed between the seasons were not reflected in a distinct trichothecene concentration in soybean seeds at maturity. Fusarium meridionale isolates showed similar pathogenicity to F. graminearum isolates but they were not able to produce this toxin in planta during the two field trials.     

Genetic variation and structure in native and invasive Solidago canadensis populations

Solidago canadensis is native to North America, but has become a noxious invasive plant in China. We know only a little about its invasion history and the effects of introductions on its genetic composition. Here, we investigated genetic variation and structure between 15 North American and 13 Chinese populations of S. canadensis using AFLP makers. Four AFLP loci suggested relatively high genetic diversity of this weed and similar genetic variation between the invasive range and the native range. Most genetic variation was within populations across two ranges, but the Chinese range had a higher degree of among‐population variation than the North American range. Multiple tests, including Bayesian assignment, UPGMA analysis, PCoA and analysis of ‘isolation by distance’, showed that the Chinese populations originated from at least two distinct native sources and that secondary introduction or dispersal should be common in China. Also, North American populations were possibly a single genetic group. Overall, S. canadensis in China was probably founded from multiple introductions and then spread through long‐distance dispersal associated with human activities. Genetic variability in the species in the invaded range appears to have favoured establishment and spread and may well provide a challenge to successful control.     

Molecular,biological and physiological characterizations of resistance to phenamacril in Fusarium graminearum

Fusarium head blight (FHB), caused by Fusarium graminearum, is one of the most devastating wheat diseases in China. Phenamacril is a novel cyanoacrylate fungicide with a unique chemical structure and specific mode of action against Fusarium spp. In this study, the molecular, biological and physiological characteristics of laboratory‐induced mutants of F. graminearum with resistance to phenamacril were investigated. Compared to the wildtype strains, the phenamacril‐resistant mutants showed obvious defects in various biological and physiological characteristics, including vegetative growth, carbon source utilization, response to oxidative and osmotic stresses, sensitivity to cell wall and cell membrane integrity inhibitors, cell membrane permeability, glycerol accumulation and pathogenicity. The phenotypes of the phenamacril‐resistant mutants exhibited many variations. Sequencing indicated that the three parental strains studied were identical, and the mutants TXR1, TXR2, BMR1, BMR2, SYR1 and SYR2 each had a single point mutation in the amino acid sequence encoded by the myosin‐5 gene (FGSG_01410). These results provide new reference information for future investigations concerning the resistance mechanism of F. graminearum to phenamacril and could offer important relevant data for the management of FHB caused by F. graminearum.     

Fusarium species associated with stalk rot and head blight of grain sorghum in Queensland and New South Wales,Australia

Historical records report Fusarium moniliforme sensu lato as the pathogen responsible for Fusarium diseases of sorghum; however, recent phylogenetic analysis has separated this complex into more than 25 species. During this study, surveys were undertaken in three major sorghum‐producing regions in eastern Australia to assess the diversity and frequency of Fusarium species associated with stalk rot‐ and head blight‐infected plants. A total of 523 isolates were collected from northern New South Wales, southern Queensland and central Queensland. Nine Fusarium species were isolated from diseased plants. Pathogenicity tests confirmed F. andiyazi and F. thapsinum were the dominant stalk rot pathogens, whilst F. thapsinum and species within the F. incarnatum–F. equiseti species complex were most frequently associated with head blight.     

Perspectives on the specific targeting of Fusarium graminearum for the development of alternative head blight treatment approaches

Diseases of agricultural crops caused by fungi have devastating economic and health effects. Fusarium head blight (FHB) is one of the most damaging diseases of wheat and other small grain cereals. FHB reduces agricultural yield while also affecting food supply and safety through deposition of toxins (mycotoxins/phytotoxins). Control of FHB growth and toxin accumulation in grains remain major challenges. While the ultimate goal in the battle against FHB is the development of resistant wheat varieties, the actual use of fully resistant plants that preclude any need for treatment with fungicides remains out of sight. Current antifungals being applied against FHB are generally azole‐based inhibitors. However, usage of these azole‐based fungicides is being complicated by the facts that these are active only during specific short‐lived developmental time periods, fungi are developing increased resistance to them and they are having significant environmental impacts. As such, there is a great need for more targeted, specific and effective antifungal agents to address the significant threat of FHB. This review provides an overview of some of the more promising fungal targets that are currently being investigated for antifungal development.     

Differential metabolic response of barley genotypes,varying in resistance,to trichothecene‐producing and ‐nonproducing (tri5−) isolates of Fusarium graminearum

Resistance of barley to Fusarium graminearum was studied using a pair each of resistant and susceptible black and yellow barley lines. The spikelets were inoculated with a trichothecene‐producing isolate, a trichothecene‐nonproducing isolate (tri5^?), or a mock solution. Spikelets were collected 72 h after inoculation and metabolites were analysed using a LC‐hybrid MS system. Metabolite abundances were used to identify the constitutive (RRC) and induced resistance‐related metabolites (RRI). The pathogen virulence factor, DON, and its plant detoxification product, DON‐3‐O‐glucoside (D3G), were also identified and designated as resistance‐indicator (RI) metabolites. The RRC, RRI and RI metabolites were putatively identified. Jasmonic acid was significantly induced in barley following inoculation with a trichothecene‐producing isolate, but not with a tri5^? isolate. The former isolate reduced the induction of both the number and amount of RR metabolites. The metabolites cinnamic acid, sinapoyl alcohol, coniferin, catechin and naringin were identified only in response to the inoculation with a tri5^? mutant. The abundances of p‐coumaric acid, coniferaldehyde and sinapaldehyde increased more in response to the tri5^? mutant than to the trichothecene‐producing isolate. The total amount of DON synthesized and its conversion to D3G varied greatly between the resistant and susceptible black barley, but not in yellow barley. Interestingly, an increase in the amount of total DON produced was associated with a decrease in the conversion of DON to D3G. The roles of RRC, RRI and RI metabolites in plant defence and their further use as potential biomarkers in screening are discussed.     

Spatial genetic structure and dispersal of the cacao pathogen Moniliophthora perniciosa in the Brazilian Amazon

Moniliophthora perniciosa is the causal agent of witches’ broom in Theobroma cacao (cacao). Three biotypes of M. perniciosa are recognized, differing in host specificity, with two causing symptoms on cacao or Solanaceae species (C‐ and S‐biotypes), and the third found growing endophytically on lianas (L‐biotype). The objectives of this study were to clarify the genetic relationship between the three biotypes, and to identify those regions in the Brazilian Amazon with the greatest genetic diversity for the C‐biotype. Phylogenetic reconstruction based on the rRNA ITS regions showed that the C‐ and S‐biotypes formed a well‐supported clade separated from the L‐biotype. Analysis of 131 isolates genotyped at 11 microsatellite loci found that S‐ and especially L‐biotypes showed a higher genetic diversity. A significant spatial genetic structure was detected for the C‐biotype populations in Amazonia for up to 137 km, suggesting ‘isolation by distance’ mode of dispersal. However, in regions containing extensive cacao plantings, C‐biotype populations were essentially ‘clonal’, as evidenced by high frequency of repeated multilocus genotypes. Among the Amazonian C‐biotype populations, Acre and West Amazon displayed the largest genotypic diversity and might be part of the centre of diversity of the fungus. The pathogen dispersal may have followed the direction of river flow downstream from Acre, Rondônia and West Amazon eastward to the rest of the Amazon valley, where cacao is not endemic. The Bahia population exhibited the lowest genotypic diversity, but high allele richness, suggesting multiple invasions, with origin assigned to Rondônia and West Amazon, possibly through isolates from the Lower Amazon population.     

Evaluation of Galium species and populations using morphological characters and molecular markers

Three Galium species are believed to be present across western Canada: Galium aparine, Galium spurium and Galium boreale. Galium spurium and G. aparine are very difficult to distinguish morphologically, which is problematic for crop consultants and weed surveyors, and could have implications for control measures. Molecular techniques could potentially make identification easier and more rapid than using chromosome counts, as is currently done. The objective of this study was to identify morphological traits and/or genetic polymorphisms capable of species differentiation. To this end, Galium seed of unknown speciation were collected from nine field populations across western Canada and, along with two reference samples of G. spurium and G. aparine, were characterised for both morphological traits and their ribosomal ITS1‐5.8S‐ITS2 genomic sequence. In addition, single nucleotide polymorphism variation within the highly conserved 5.8S ribosomal RNA gene was identified that could consistently differentiate Galium species. Sequence analysis of the ITS1‐5.8S‐ITS2 region of field collections from western Canada indicated that all samples were G. spurium and all were highly related to each other. These results were supported by a distinct lack of variation in morphological traits, as nearly all plant traits measured did not differ between populations. This suggests that all sampled populations, and perhaps most of the Galium populations across western Canada, are derived from a single species, G. spurium.     

Genetic diversity and structure of Hemileia vastatrix populations on Coffea spp.

Coffee leaf rust is the most limiting disease for coffee cultivation in Brazil. Despite its importance, relatively little is known about the genetic diversity of Hemileia vastatrix, the rust causal agent. In this work, the DNA from 112 monopustule isolates from different geographic locations and coffee genotypes were analysed by amplified fragment length polymorphisms (AFLP). The objectives were to assess the influence of the host and geographic origin on the diversity and population differentiation in H. vastatrix. The fungal population showed a low level of genotypic diversity. Gene diversity (h) was 0·027 and the hypothesis of random mating in the total population was rejected, but evidence for recombination was found for two subpopulations (São Paulo and Paraná). The analysis of molecular variance revealed that 90% of the genetic distribution of the pathogen occurs among isolates within the subpopulation (states or host of origin). There was no correlation between geographic and genetic distance (r = ?0·024, P = 0·74), which together with the high number of migrants and the low degree of differentiation in populations of H. vastatrix, is consistent with the fact that the inoculum is probably easily dispersed by wind over long distances, allowing dispersal of the pathogen among coffee growing areas in Brazil. Therefore, it is difficult to predict the durability of resistant sources to coffee rust. The recommendation for the breeding programmes is thus to incorporate multigenic resistance as a control strategy.     

Evidence for a new introduction of the pitch canker fungus Fusarium circinatum in South Africa

Fusarium circinatum causes pitch canker of Pinus species in many parts of the world. The fungus was first recorded in South Africa in 1990 as a pathogen of P. patula seedlings and emerged later as a pathogen of established plantation trees, especially P. radiata in the Western Cape Province (WCP). In this study the population biology of F. circinatum in the WCP was explored. The aim was to determine the possible origin and reproductive mode of the pathogen, with the ultimate intention of informing disease management strategies in the region. Vegetative compatibility assays, sexual mating studies and amplified fragment length polymorphism analyses were used. For comparative purposes, an isolate collection obtained from diseased P. radiata seedlings in a commercial nursery in the region, as well as a set of isolates from commercial seedling nurseries in the central and northern parts of South Africa, were included. The results showed that the WCP population of F. circinatum employs a predominantly asexual mode of reproduction and that it is highly differentiated from populations of the fungus elsewhere in South Africa. However, limited genetic structure was found within the respective WCP isolate collections. Overall these findings suggest that pitch canker in the WCP originates from one or more separate introductions of the pathogen and that its movement in the region is not restricted. More effective strategies are thus required to limit and manage the effects of F. circinatum in plantations in this region of South Africa.     

Genetic variation of the invasive Campuloclinium macrocephalum,Asteraceae in South Africa,inferred from molecular markers

Campuloclinium macrocephalum is native to Central and South America, but is a highly invasive weed in South Africa, where it is commonly known as the ‘pompom weed’. It is targeted for biological control, the success of which will depend on host specificity and biotype compatibility to its full genetic diversity in South Africa. We investigated the genetic diversity and phylogeography of 52 specimens from across South Africa, 14 from Argentina and three from Brazil using nuclear ribosomal ITS regions. We further explored the AFLP marker diversity in 54 South African, 25 Argentine and three Brazilian specimens. Maximum parsimony analysis of the ITS sequence data produced an unresolved phylogeny. However, three haplotypes were recognised via network analysis. All South African, one Brazilian and all bar one of the Argentine individuals shared a single haplotype. AFLP analyses generated two genetic clusters with a low net nucleotide distance of 0.115 and further revealed that most plants were a mixture of alleles from these two genetic clusters. Although there was a significant genetic variation among the populations, genetic differentiation and mean heterozygosity were low, suggesting that clonal reproduction may be occurring. The current South African populations may therefore be clonal products with different proportions of genetic admixture introduced more than once. The original point of entry appears to be Gauteng, South Africa. Long‐distance dispersal appears to have played a major role in its spread across South Africa. Candidate Argentine biological control agents should therefore be effective on C. macrocephalum in South Africa.     

Crop type and altitude are the main drivers of species composition of arable weed vegetation in Tajikistan

The influence of broad‐scale environmental factors on the species composition of segetal weed communities in Tajikistan was investigated. The research was conducted throughout the country, analysing plots of root crops as well as cereals from all phytogeographical regions of Tajikistan, with the exception of the eastern Pamir. The study was based on 440 phytosociological relevées sampled between 2009 and 2013 and analysed using direct and indirect ordinations. A set of environmental variables was obtained for each plot: altitude, mean annual precipitation, mean annual temperature, pH, crop type, longitude, latitude and date of sampling (seasonality). Crop type was the major factor determining species composition, which was related to the different farming practices in roots and cereals. Major changes in weed species composition were also driven by altitude and correlated temperatures. The seasonality of weed communities also had a considerable effect on weed community structure. Samples from spring differed significantly from samples from summer and late summer. Only longitude and soil pH were found to be insignificant. The complete set of environmental variables used in a canonical correspondence analysis explained 8.3% of the species variation in sampled plots. The study showed that despite the very diverse climatic conditions and low intensity of agriculture in Tajikistan, agrocoenoses respond mainly to the type of cultivation practised in arable land.     

Nursery‐linked plantation outbreaks and evidence for multiple introductions of the pitch canker pathogen Fusarium circinatum into South Africa

In recent years, Pinus plantation forestry has been significantly hampered by outbreaks of pitch canker caused by the fungus Fusarium circinatum. This study investigated the role of Pinus host, geographic origin and reproductive mode in structuring the F. circinatum populations in plantations. For this purpose, 159 isolates originating from diseased plantation trees in the Western and Eastern Cape Provinces of South Africa were genotyped using 10 microsatellite markers. Analyses of these data revealed 30 multilocus haplotypes and that the populations were distinct based on geographic origin as well as host. However, shared haplotypes were observed between populations, showing that these populations are connected, possibly through the movement of haplotypes. A second aim was to determine whether the genetic variation found in these populations of the fungus could be attributed to outbreaks of the seedling disease caused by this pathogen in Pinus nurseries. To achieve this goal, an additional set of 43 isolates originating from pine seedling nurseries was genotyped and analysed. The results showed that the populations of F. circinatum in plantations most probably originated from the nursery outbreaks that occurred prior to the plantation outbreak. Inferences regarding reproductive mode further showed that sexual reproduction has little impact on the genetic makeup of the F. circinatum populations and that they primarily reproduce asexually. Overall, the results of this study showed that the F. circinatum diversity in South Africa has arisen due to multiple introductions of the pathogen and is not due to sexual reproduction.     

Species of the Colletotrichum gloeosporioides and C. boninense complexes associated with olive anthracnose

The taxonomic status of Colletotrichum gloeosporioides sensu lato (s.l.) associated with olive anthracnose is still undetermined and the pathogenic ability of this species complex is controversial. In the present study, isolates obtained from olive and provisionally identified as C. gloeosporioides s.l. on the basis of morphological and cultural features were reclassified using ITS and TUB2 as DNA barcode markers and referred to seven distinct species, recently separated within C. gloeosporioides (C. aenigma, C. gloeosporioides sensu stricto (s.s.), C. kahawae, C. queenslandicum, C. siamense and C. theobromicola) and C. boninense (C. karstii) species complexes. Furthermore, isolates of C. kahawae were ascribed to the subspecies ciggaro by analysing the GS gene. A single isolate, not in either of these two species complexes, was not identified at the species level. In pathogenicity tests on detached olive drupes some of these species, including C. aenigma, C. kahawae subsp. ciggaro, C. queenslandicum, C. siamense and C. karstii, were shown to be weakly pathogenic. Moreover, they were found very sporadically on olive. In contrast, some isolates of C. gloeosporioides s.s. and isolates of C. theobromicola proved to be virulent on both green and ripening olives. This study gives a better insight into both the aetiology and the epidemiology of olive anthracnose and might have implications for biosecurity and quarantine because C. theobromicola has never been reported in major European olive‐producing countries.     

Spatiotemporal distribution of Ascochyta pinodes and Ascochyta pinodella during the winter growing season in France

Ascochyta blight of pea is caused by four related fungi, Ascochyta pisi, Phoma koolunga, Ascochyta pinodes and Ascochyta pinodella. The latter two taxa appear to be much more common and economically significant worldwide but the relative impact of each fungus on ascochyta blight epidemics is not well understood. To study the spatiotemporal distribution of A. pinodes and A. pinodella infecting pea in France, 368 isolates were sampled monthly, from February to May, at three locations (Rennes, Boigneville and Dijon) and molecular markers were used to genotype isolates. The aggressiveness of isolates from the fourth sampling date was estimated using a detached leaf assay on the winter cultivar Enduro. Disease was low during the sampling period as climatic conditions were generally not conducive to disease development (cold temperature, low rainfall). Population genetic analysis showed that 99% of the observed variation could be attributed to variation within populations compared to only 1% among populations. Both species were observed in each location, although A. pinodella was observed at a lower frequency (6–32%). Moreover, results showed that both species could develop on different nodes of the plant. Significant differences in aggressiveness were observed between species and among isolates within species with A. pinodes isolates being significantly more aggressive on average than A.  pinodella isolates. These results emphasize the necessity to study the components of disease complexes in order to understand the impact of pathogen species interactions on disease and yield reduction as well as the dynamics of disease epidemics during the cropping season.