Septoria Leaf Spot of Banana: A Newly Discovered Disease Caused by Mycosphaerella eumusae (Anamorph Septoria eumusae)

ABSTRACT A previously undescribed leaf spot disease of banana has been discovered in southern and Southeast Asia. The fungus identified as the causal agent of this leaf spot has a Mycosphaerella teleomorph stage and a Septoria anamorph stage. Isolation and reinoculation of the fungus to banana reproduced symptoms and confirmed its pathogenicity. Phylogenic analysis based on sequences of the internal transcribed spacer and 5.8S ribosomal DNA regions from the different leaf spot pathogens of bananas was consistent with the definition of a new species. M. eumusae (anamorph S. eumusae) is the name proposed for the causal agent and Septoria leaf spot as the name for the disease. The presence of the pathogen has been confirmed in leaf specimens from southern India, Sri Lanka, Thailand, Malaysia, Vietnam, Mauritius, and Nigeria.     

The Occurrence of Mycosphaerella graminicola and its Anamorph Septoria tritici in Winter Wheat during the Growing Season

The disease septoria tritici blotch of wheat is initiated by ascospores of the teleomorph Mycosphaerella graminicola or pycnidiospores of the anamorph Septoria tritici. We report for the first time the presence of the teleomorph, M. graminicola, in Denmark. With the objective of elucidating the importance of the teleomorph for the development of septoria tritici blotch, data on the occurrence of fruit bodies of the anamorph (pycnidia) and the teleomorph (pseudothecia) stages were collected over three growing seasons. Pseudothecia were present in the springs, however, high numbers of pseudothecia compared to pycnidia were not observed until July, too late to influence the epidemic. On an individual leaf layer, pycnidia were observed well before pseudothecia. As the leaves aged, progressively higher proportions of fruit bodies were observed to be pseudothecia. The period from the appearance of pycnidia to detection of pseudothecia was estimated as 29–53 days. At harvest, high proportions of sporulating fruit bodies in the crop were pseudothecia, suggesting that the primary source of inoculum for new emerging wheat crops in autumn is likely to be ascospores.     

Genetic Structure of Mycosphaerella graminicola Populations from Iran, Argentina and Australia

Restriction fragment length polymorphism (RFLP) markers were used to assess the genetic structure of populations of Mycosphaerella graminicola collected from wheat fields. A total of 585 isolates representing 10 field populations were sampled from Iran, Argentina and Australia. The genetic structure of M. graminicola populations from Iran and Argentina is described for the first time. Results were compared to previously investigated populations from Israel, Uruguay and Australia. Populations from Iran exhibited high clonality and low gene diversity, suggesting an inoculation event. Populations from uninoculated fields in Argentina had gene and genotype diversities similar to previously described European and North American populations. Genotype diversity was high for populations from Australia and tests for multilocus associations were consistent with sexual recombination in these populations. Gene diversity was low and fixed alleles were found for several loci. These findings are consistent with a relatively small founding population for Australia. These 10 new populations were integrated into a genetic distance comparison with 13 global populations that were characterized earlier.     

Partial Resistance to Septoria Tritici Blotch (Mycosphaerella graminicola) in Wheat Cultivars Arina and Riband

ABSTRACT Partial resistance to Septoria tritici blotch (STB) and its inheritance were investigated in a doubled-haploid population of a cross between cvs. Arina and Riband. The former has good partial resistance whereas the latter is susceptible. In adult plant trials in polytunnels, STB disease scores were negatively correlated with heading date. Resistance was not specific to any of the three fungal isolates used in these tests. A quantitative trait locus (QTL) for partial resistance to STB was identified in Riband on chromosome 6B and is named QStb.psr-6B-1. No QTL controlling a major part of the Arina resistance was identified, suggesting that its resistance may be dispersed and polygenic. There was no correlation between the lines' mean disease scores at the seedling and adult stages, implying that partial resistance to STB is developmentally regulated. Seedling resistance to the isolate IPO323 was isolate-specific and controlled by a single gene in Arina, probably allelic with the Stb6 gene in cv. Flame that confers resistance to the same isolate. The implications of these results for wheat breeding programs are discussed.     

The Genetic Structure of Australian Populations of Mycosphaerella musicola Suggests Restricted Gene Flow at the Continental Scale

ABSTRACT Mycosphaerella musicola causes Sigatoka disease of banana and is endemic to Australia. The population genetic structure of M. musicola in Australia was examined by applying single-copy restriction fragment length polymorphism probes to hierarchically sampled populations collected along the Australian east coast. The 363 isolates studied were from 16 plantations at 12 sites in four different regions, and comprised 11 populations. These populations displayed moderate levels of gene diversity (H = 0.142 to 0.369) and similar levels of genotypic richness and evenness. Populations were dominated by unique genotypes, but isolates sharing the same genotype (putative clones) were detected. Genotype distribution was highly localized within each population, and the majority of putative clones were detected for isolates sampled from different sporodochia in the same lesion or different lesions on a plant. Multilocus gametic disequilibrium tests provided further evidence of a degree of clonality within the populations at the plant scale. A complex pattern of population differentiation was detected for M. musicola in Australia. Populations sampled from plantations outside the two major production areas were genetically very different to all other populations. Differentiation was much lower between populations of the two major production areas, despite their geographic separation of over 1,000 km. These results suggest low gene flow at the continental scale due to limited spore dispersal and the movement of infected plant material.     

Genotypic Diversity of the Wheat Leaf Blotch Pathogen Mycosphaerella Graminicola (anamorph) Septoria Tritici in Germany

The population structure and genotypic diversity of Mycosphaerella graminicola from six natural field populations in Germany were studied with molecular markers. To reveal the potential effects of plant host resistance on the pathogen population, hierarchical samples were taken from susceptible and resistant cultivars. A total of 203 single spore isolates was subjected to molecular marker analysis using the amplified fragment length polymorphism technique (AFLP). Among the 203 isolates analyzed, 142 different multilocus haplotypes (MLH) were identified revealing a high degree of genotypic diversity of the M. graminicola population. On average, a F _ST value of 0.04 was found, indicating a low genetic differentiation with only 4% of the genetic variation between the local populations but leaving 96% of the genetic variation within the populations. According to the low F _ST value, a high migration rate of Nm 12 was found. The observed high within-population diversity, and the significant migration between populations, prevented genetic isolation and differentiation of putative geographically separated populations. Furthermore, plant host resistance had no obvious effect on the population structure and diversity of M. graminicola. Genotypic variability can be attributed to sexual recombination which appears to have a considerably larger influence on the population structure. Gene flow on this scale could have significant implications for plant breeding and fungicide spraying programmes.     

Factors determining the severity of epidemics of Mycosphaerella graminicola (Septoria tritici) on winter wheat in the UK

Epidemics of disease caused by Septoria tritici were studied in detail in 11 crops of winter wheat cv. Longbow over 4 years. Serious damage to the uppermost two leaf layers was caused by splash-borne infection from lower in the crop early in the life of the leaves, followed by one or rarely two cycles of multiplication within a leaf layer. Infection conditions rarely limited damage, even in a dry year; the timing and, to a lesser extent, amount of initial inoculum movement to an upper leaf layer was of greater importance. Timing of initial infection was determined by when rain splash occurred in relation to emergence of a leaf layer. Occurrence of infections soon after a leaf layer started to emerge allowed more time for multiplication of disease within that layer. These infections tended to be more severe because the leaves were closer to inoculum sources within the crop. Slight differences in phenology between locations explain why initially random disease distributions sometimes become aggregated. Early-sown crops are at greater risk because they mature more slowly, allowing more disease multiplication and better transfer between leaf layers.     

Phytophthora infestans Populations from Tomato and Potato in North Carolina Differ in Genetic Diversity and Structure

ABSTRACT Phytophthora infestans causes a destructive disease on tomato and potato. In North Carolina (NC) potatoes are mostly grown in the east, whereas tomatoes are grown in the mountainous areas in the western part of the state. Five genotypes of P. infestans were identified from 93 and 157 isolates collected from tomato and potato over a 5 year period between 1993 and 1998. All isolates collected from potato in eastern NC were the US-8 genotype, whereas only a single isolate was the US-1 genotype. Tuber blight was found on immature daughter tubers in a single field in 1997, however infection on mature tubers was not observed. Within potato fields, a range of sensitivity to metalaxyl was observed among isolates but all were either intermediate or highly resistant to the fungicide. In contrast, isolates from tomatoes included previously reported US-7 and US-8 genotypes and two new genotypes called US-18 and US-19 (A2 mating type, allozyme genotype Gpi 100/100 and Pep 92/100). These genotypes had unique restriction fragment length polymorphism banding patterns, were sensitive to metalaxyl, and have not been reported elsewhere. All genotypes, with the exception of the US-1, were the Ia mitochondrial haplotype. Thus, isolates of P. infestans from tomato were more genetically diverse over time in NC than those from potato and include two new genotypes that are sensitive to metalaxyl.     

Population Genetic Structure of Septoria passerinii in Northern Great Plains Barley

ABSTRACT The genetic structure of Septoria passerinii from nine field populations was examined at several scales (within lesions, among lesions in a leaf, among leaves in a field, and among fields in North Dakota and western Minnesota) by using amplified fragment length polymorphism (AFLP) markers. A total of 390 isolates were sampled from seven barley fields located in North Dakota and two barley fields located nearby in western Minnesota in 2003 and 2004. Based on 57 polymorphic AFLP markers, AFLP DNA fingerprints identified 176 different genotypes among 390 (non-clone-corrected) isolates in nine different fields. In two intensively sampled sites, ND16 (Williston, ND) and ND17 (Langdon, ND), only one to four different genotypes were found within a lesion. A higher level of genetic and genotypic diversity was found within a leaf in which six to nine different genotypes were found from lesions on a leaf. The genetic diversity within a leaf was similar to the genetic diversity within a field. The average genetic diversity (H) within a field across all AFLP loci was approximately 0.3, except at site ND12 (Carrington, ND) where it was 0.16. Genotypic diversity was high in all populations, and with the exception of ND15 (Rothsay, MN), very low multilocus linkage disequilibrium values ( r(d)) were found in all populations. The population differentiation, G(ST), was relatively high (G(ST) = 0.238) among the nine populations due to the high G(ST) in ND12, ND14 (Twin Valley, MN), and ND15. Population differentiation without those three populations was 0.09. A lack of correlation between geographical distance and genetic distance was found, suggesting the potential for a high level of gene flow between different geographical regions. The population genetic structure described in this study for S. passerinii in North Dakota and western Minnesota is consistent with that of a sexually reproducing fungus.     

Influence of crop growth and structure on the risk of epidemics by Mycosphaerella graminicola (Septoria tritici) in winter wheat

Generally, it is recognized that inocula of Septoria tritici present on the basal leaves of winter wheat crops are spread towards the top of the canopy by splashy rainfall. This mechanism of inoculum dispersal is commonly accepted to be a key limit on disease progression. Therefore, attempts to forecast epidemics of S. tritici often quantify rainfall by some means, but largely ignore measurement of pathogen and host variables. In the present study, we show that new wheat leaves emerge initially at a height below established leaves that can contain sporulating lesions of S. tritici . This presents the possibility of horizontal inoculum transfer, even without splashy rainfall. The extent and duration of overlap between emergent and established leaves was found to differ considerably with cultivar and sowing date. Nitrogen application had little effect on overlap, because differences in crop phenology, e.g. leaf area and nodal length, were relative. However, estimates of raindrop penetration to the base of crop canopies suggested that vertical movement of inoculum is affected by nitrogen application. Crops receiving more nitrogen are denser, and therefore less rainfall reaches the base of the canopy. The interactions between crop and pathogen development are discussed with reference to the implications for predicting disease risk. In particular, cultivar traits that promote disease escape are quantified.     

A detached seedling leaf technique to study resistance to Mycosphaerella graminicola (anamorph Septoria tritici) in wheat

A detached seedling leaf technique was developed to screen for resistance to septoria tritici blotch of wheat and to detect specific interactions between cultivars and isolates. Wheat seedlings were inoculated with spore suspensions of Mycosphaerella graminicola . Detached primary leaves were then placed in a clear plastic box such that their cut ends were sandwiched between layers of agar containing benzimidazole, with a gap below the middle of the leaves. Mean levels of disease were affected by light and temperature, and also by the concentration of benzimidazole, such that higher concentrations resulted in less disease. Second leaves were more susceptible than seedling primary leaves. However, none of these factors affected ranking of disease among cultivars or cultivar-by-isolate interactions. Kavkaz–K4500 1.6.a.4, Synthetic 6x and Triticum macha showed specific susceptibility and resistance to different isolates. The detached leaf technique could be a useful complement to field trials and an alternative to whole seedling assays in assessing cultivar resistance and investigating the genetics of the host–pathogen interaction.     

Interacting effects of interrupted humid periods and light on infection of wheat leaves by Mycosphaerella graminicola (Septoria tritici)

Septoria tritici conidia require a long period at 100%, humidity to infect wheat plants successfully. When periods of 100% relative humidity were interrupted by up to 48 h at 75% relative humidity, infection by S. tritici on winter wheat cvs Longbow and Avalon was only slightly reduced. Breaks at 50% relative humidity had larger effects, but still allowed infection to occur. Infection was reduced more by a break in humidity which began in the light, but previous and subsequent light regimes strongly influenced the final outcome in all cases. When a wet period was interrupted twice, the two dry breaks interacted strongly under some circumstances; in these cases the probability of infection was much greater than expected from multiplying the effects of the single breaks. Under other conditions two breaks had more nearly multiplicative effects, or interacted to reduce the probability of infection below that expected from the individual breaks. There was evidence that light reduced the probability of successful infection earls in the infection process, but stimulated it later. The data were consistent with infection occurring faster on cv. Longbow, but otherwise being affected in qualitatively similar ways on both cultivars. The complexity of the effects and their interactions suggests that it will not be possible to find a simple set of infection conditions to apply in the field. The data rule out a number of simple mathematical models of the infection process.     

Sensitivity of Mycosphaerella graminicola (anamorph: Septoria tritici) to DMI fungicides across Europe and impact on field performance

Since the occurrence and spread of resistance to quinone outside inhibitors (QoI) in Mycosphaerella graminicola in the early 2000s in Europe, demethylation inhibitors (DMIs) form the backbone for control of Septoria leaf blotch. European monitoring studies, carried out by various research institutes and DMI manufacturers, have shown a shift of the European M. graminicola population towards increased ED₅₀ values for DMI fungicides. Populations of M. graminicola consist of very heterogeneous isolates within a region, and even within a field, in terms of DMI sensitivity. Sensitivity to DMIs is influenced by the haplotype of CYP51, the target of DMIs. New CYP51‐haplotypes have emerged and the frequency of less sensitive haplotypes in Europe has increased in recent years. Studies with efflux transporter inhibitors showed that not only CYP51, but also enhanced efflux, may play a role in the DMI sensitivity response. Sensitivity studies with 5 DMIs registered for Septoria leaf blotch control indicated that sensitivity of isolates to the 5 DMIs is heterogeneous and the overall correlation of sensitivity to the different DMIs is poor. A key requirement for sustainable control and resistance management of Septoria leaf blotch is therefore the continued availability of different DMIs.     

Airborne inoculum as a major source of Septoria tritici (Mycosphaerella graminicola) infections in winter wheat crops in the UK

The pattern and extent of primary infection by Septoria tritici were compared over a period of 3 years in winter wheat grown at sites with differing histories and from seed stocks obtained in different countries, in the open, under airtight cover and in sterilized soil. Only the airtight cover altered the number of lesions found, substantially reducing it. Lesions were evenly distributed. Lesions were found throughout the autumn and occasionally in the winter and spring on wheat seedlings exposed in trays to the open air for periods of 1 week, then given good conditions for infection to occur. This was true even at a site 0.4 km distant from wheat residues. The results show that the main source of primary infection of winter wheat in these experiments was evenly dispersed and airborne. It probably consisted mainly of ascospores of Mycosphaerella graminicola.     

A Model of the Effect of Pseudothecia on Genetic Recombination and Epidemic Development in Populations of Mycosphaerella graminicola

ABSTRACT It is generally agreed that ascospores are the origin of primary infections for the disease septoria tritici blotch of wheat caused by the fungus Mycosphaerella graminicola (anamorph Septoria tritici). The epidemic during the growing season was previously ascribed to the asexual pycni-diospores dispersed over short distances by rain splash, but recent observations suggest that the airborne ascospores also may play a role. As a consequence, the composition of the pathogen population over the growing season may change through genetic recombination. In an attempt to resolve the relative importance of the two spore types to the epidemic over the growing season, a model simulating disease caused by both types of spores was constructed and analyzed. The conclusion from the analysis of this model is that sexual recombination will affect the genetic composition of the population during a growing season. A considerable proportion of spores released at the end of the growing season may be sexual descendants of the initial population. However, ascospores are unlikely to affect the severity of the epidemic during the growing season. This is due to the much longer latent period for pseudothecia compared with pycnidia, resulting in ascospores being produced too late to influence the epidemic.     

Genetic Structure of Cronartium ribicola Populations in Eastern Canada

ABSTRACT The genetic structure of populations of Cronartium ribicola was studied by sampling nine populations from five provinces in eastern Canada and generating DNA profiles using nine random amplified polymorphic DNA markers. Most of the total gene diversity (H(t) = 0.386) was present within populations (H(w) = 0.370), resulting in a low level of genetic differentiation among populations in northeastern North America (F(st) = 0.062). A hierarchical analysis of genetic structure using an analysis of molecular variance (AMOVA) revealed no statistically significant genetic differentiation among provinces or among regions. Yet, genetic differentiation among populations within regions or provinces was small (AMOVA phi(st) = 0.078) but statistically significant (P < 0.001) and was several orders of magnitude larger than differentiation among provinces. This is consistent with a scenario of subpopulations within a metapopulation, in which random drift following migration and new colonization are major evolutionary forces. A phenetic analysis using genetic distances revealed no apparent correlation between genetic distance and the province of origin of the populations. The hypothesis of isolation-by-distance in the eastern populations of C. ribicola was rejected by computing Mantel correlation coefficients between genetic and geographic distance matrices (P > 0.05). These results show that eastern Canadian provinces are part of the same white pine blister rust epidemiological unit. Nursery distribution systems are controlled provincially, with virtually no seedling movement among provinces; therefore, infected nursery material may not play an important role in the dissemination of this disease. Long-distance spore dispersal across provincial boundaries appears to be an epidemiologically important factor for this pathogen.     

Genetic Structure of Peruvian Populations of Phytophthora infestans

ABSTRACT Isolates of the late blight pathogen Phytophthora infestans (n = 327) from the central to southern Peruvian Andes were systematically collected in 1997 to 1999 and analyzed to determine the pathogen's population structure at its host's center of diversity. No isolates of the A2 mating type were detected. Cluster analysis of DNA fingerprinting data indicated that the collection consisted of five major groups that were interpreted to be clonal lineages. Two of the lineages (US-1 and EC-1) have been previously described, and three (PE-3, 5, and 6) are described here for the first time. Collections from three areas in the central Peruvian Andes, including two key sites used in an international potato breeding program, consisted of isolates of the EC-1 lineage, which has been reported to dominate the pathogen population in Andean countries to the north of Peru. The collections from Cusco and Puno were more diverse. More than one lineage was detected in 10 of the 20 fields sampled in Cusco. Data on virulence, metalaxyl sensitivity, and band data for allozymes, mitochondrial DNA, and ipiB1 suggested that PE-3 may have been produced through recombination events between US-1 and EC-1. Restriction fragment length polymorphism and amplified fragment length polymorphism marker data were not consistent with this hypothesis.     

Measuring Immigration and Sexual Reproduction in Field Populations of Mycosphaerella graminicola

ABSTRACT A field experiment was conducted to determine the relative contributions of immigration and sexual reproduction to the genetic structure of Mycosphaerella graminicola populations during the course of an epidemic. The genetic structure of M. graminicola populations sampled from wheat plots inoculated artificially with 10 isolates was compared with control plots infected naturally by airborne ascospores. Restriction fragment length polymorphisms (RFLPs) were used to test the randomness of associations among loci, and DNA fingerprints were used to identify clones. All isolates in the control plots had unique genotypes and RFLP loci were at gametic equilibrium, findings consistent with random mating. The proportion of isolates in the inoculated plots with DNA fingerprints that differed from the 10 inoculated isolates increased from 3% in the early to 39 and 34% in the mid- and late season, respectively. The degree of gametic disequilibrium was higher in the mid-season than in the late-season population. By the end of the growing season, we estimate that 66% of the isolates in the inoculated plots were asexual progeny of the 10 inoculated isolates, 10% were immigrants, and 24% were sexual recombinants. The proportion of infections caused by ascospores increased over the growing season.     

Genetic Structure of Setosphaeria turcica Populations in Tropical and Temperate Climates

ABSTRACT Northern leaf blight, caused by Setosphaeria turcica, is a serious disease of maize in temperate and tropical environments. To examine the pathogen's population structure, we analyzed 264 isolates from four different continents with 70 random amplified polymorphic DNA markers and determined their mating types. Tropical populations (from Kenya, Mexico, and southern China) had an extremely high genotypic diversity, no or only weak gametic phase disequilibrium, and an even distribution of the two mating types, indicating frequent sexual recombination. Temperate populations (from Europe and northern China) had a much lower genotypic diversity, strong gametic phase disequilibrium, and an uneven distribution of mating types, indicating that sexual recombination has been rare. Populations in different continents were genetically isolated. They shared no haplotypes and carried several "private" alleles. The number of migrants between continents and between regions (between northern and southern China, western and central Kenya, and Europe west and east of the Alps) was estimated to be less than one per generation. Multivariate statistics suggested a greater relatedness of populations from the same continents than from different continents. Within agroecological zones, migration must be extensive. The potential within populations of S. turcica for adaptation should be regarded as very high, especially in tropical climates.     

Barrier to Gene Flow Between Eastern and Western Populations of Cronartium ribicola in North America

ABSTRACT The population structure of Cronartium ribicola from eastern and western North America was studied to test the null hypothesis that populations are panmictic across the continent. Random amplified polymorphic DNA markers previously characterized in eastern populations were mostly fixed in western populations, yielding high levels of genetic differentiation between eastern and western populations (phi(st) = 0.55; theta = 0.36; P < 0.001). An unweighted pair-group method, arithmetic mean dendro-gram based on genetic distances separated the four eastern and four western populations into two distinct clusters along geographic lines. Similarly, a principal component analysis using marker frequency yielded one cluster of eastern populations and a second cluster of western populations. The population from New Mexico was clearly within the western cluster in both analyses, confirming the western origin of this recent introduction. This population was completely fixed (H(j) = 0.000; n = 45) at all loci suggesting a severe recent population bottleneck. Genetic distances were low among populations of western North America (0.00 to 0.02) and among eastern populations (0.00 to 0.02), indicating a very similar genetic composition. In contrast, genetic distances between eastern and western populations were large, and all were significantly different from 0 (0.07 to 0.19; P < 0.001). Indirect estimates of migration were high among western populations, including the number of migrants among pairs of populations (Nm > 1) between New Mexico and British Columbia populations, but were smaller than one migrant per generation between eastern and western populations. These results suggest the presence of a barrier to gene flow between C. ribicola populations from eastern and western North America.