Seasonal Variation in Sporulation of Phytophthora infestans

Sporulation ability of two isolates of Phytophthora infestans maintained on potato tuber slices of a susceptible cv. ‘Bintje’ and on-rye agar medium was studied for four years (1981–1984). This feature of the fungus was shown to vary in particular seasons during the year. Significantly higher sporulation density per cm² of aerial mycelium on potato tuber slices was observed in winter and late autumn while significantly lower sporulation was found in spring. Similar tendencies were observed when one of the isolates was cultivated on rye agar medium under controlled conditions. Positive correlation was found between sporulation patterns of isolates of the fungus maintained on rye agar and on tuber slices. Hypothesis has been proposed that these changes are due, to a biorhythm in the fungus.     

Seasonal variation of arbuscular mycorrhizal fungi in temperate grasslands along a wide hydrologic gradient

We studied seasonal variation in population attributes of arbuscular mycorrhizal (AM) fungi over 2 years in four sites of temperate grasslands of the Argentinean Flooding Pampas. The sites represent a wide range of soil conditions, hydrologic gradients, and floristic composition. Lotus glaber, a perennial herbaceous legume naturalised in the Flooding Pampas, was dominant at the four plant community sites. Its roots were highly colonised by AM fungi. Temporal variations in spore density, spore type, AM root colonisation, floristic composition and soil chemical characteristics occurred in each site and were different among sites. The duration of flooding had no effect on spore density but depressed AM root colonisation. Eleven different types of spores were recognized and four were identified. Two species dominated at the four sites: Glomus fasciculatum and Glomus intraradices. Spore density was highest in summer (dry season) and lowest in winter (wet season) with intermediate values in autumn and spring. Colonisation of L. glaber roots was highest in summer or spring and lowest in winter or autumn. The relative density of G. fasciculatum and G. intraradices versus Glomus sp. and Acaulospora sp. had distinctive seasonal peaks. These seasonal peaks occurred at all four sites, suggesting differences among AM fungus species with respect to the seasonality of sporulation. Spore density and AM root colonisation when measured at any one time were poorly related to each other. However, spore density was significantly correlated with root colonisation 3 months before, suggesting that high colonisation in one season precedes high sporulation in the next season.     

Phenotypic markers of Phytophthora infestans

Abstract

Phytophthora infestans is one of the most destructive pathogens of potato and causal agents of notorious disease late blight. Different chemicals are used to control the pathogen of late blight but the most commonly used is metalaxyl; its extensive use of has caused decreased sensitivity in the P. infestans population. The metalaxyl sensitivity of the Pakistani population of P. infestans is investigated in the present study. For this purpose, 178 isolates of P. infestans were obtained from the lesions of diseased potato leaves and stems, and samples were collected from the different potato-growing areas of Pakistan, where late blight is a problem. Sensitivity of the isolates of P. infestans was investigated by metalaxyl sensitivity test and with the help of test isolates were divided into three categories, i.e. sensitive, intermediate and resistant, based on their Co-efficient of mycelial growth inhibition (CMGI) values. During the study, highest percentage (50.17%) of resistant isolates was observed in the population of Punjab (zone 2), whereas the lower percentage (33.33%) was observed in the population of Swat valley (zone 6b). In the present study, it was discovered that P. infestans late blight-causing fungus has adopted more resistance against metalaxyl because of its wide use.     

Aggressiveness variation in Plasmopara halstedii and its alternation with non-race-specific resistance in sunflower

Aggressiveness variation and its alternation with non-race specific resistance in sunflower were studied in 19 Plasmopara halstedii isolates belonging to several races. Regarding aggressiveness criteria, percentage infection, latent period, sporulation density and dwarfing, on two sunflower inbred lines showing different levels of non-race specific resistance resistance FU and BT, there were significant differences in aggressiveness for P. halstedii isolates. The index of aggressiveness varied between 9.4 and 31.4. The inbred line BT, rather susceptible in the field, showed a higher percentage infection, a higher sporulation density, a shorter latent period and less reduced hypocotyl length than inbred line FU, which showed a greater resistance in the field. Percentage infection on FU was 1.4% less than BT, latent period on BT was 12.4% less than FU, sporulation density on FU was 22.3% less than BT and reduced hypocotyl length on BT was 15.3% less than FU. Consequently, it seems that the criteria as latent period, sporulation density and reduction of hypocotyl length could be used to measure non-race specific resistance in sunflower to P. halstedii under controlled conditions.     

Spectral filters for the control of Botrytis cinerea

Experiments performed in vitro examined the sporulation of Botrytis cinerea (grey mould) under different spectral distributions. Eighty‐three isolates, taken from plants of primula (Primula vulgaris) at different locations throughout the UK, were incubated in the dark, with visible light only and visible plus near‐ultraviolet (nUV) light. On average, compared to isolates not exposed to nUV, sporulation was increased 54‐fold following illumination with nUV light. No isolates showed complete insensitivity to near ultraviolet. New polyethylene materials with different optical properties were then tested on two typical isolates. A film which removed nUV up to 405 nm, compared to a film with nUV absorption up to 384 nm, resulted in the lowest production of conidia (by 5‐fold). The former film was used to clad horticultural polyethylene tunnels in which crops of P. vulgaris and strawberry were grown for two seasons and the incidence of B. cinerea assessed throughout the growth of the crops. The incidence of infection on the P. vulgaris and strawberries was reduced by c. 50% and c. 26% respectively with the nUV blocking film compared to a standard film. The results are discussed in terms of the potential of spectral filters as a novel means of grey mould control in greenhouse‐produced crops.     

Use of a novel two-stage cultivation method to determine the effects of environmental factors on the growth and sporulation of several biocontrol fungi

To supply essential information for improving mass production and biocontrol efficacy, two-stage cultivation on agar plates was used to evaluate the environmental conditions affecting mycelial growth and sporulation of seven biocontrol fungi. Maximum growth and sporulation occurred on acid media for Paecilomyces (Pa.) lilacinus IPC-P, Pochonia (Po.) chlamydosporia HSY-12-14, and Lecanicillium lecanii CA-1-G, and on alkaline media for Metarhizium anisopliae isolates. All fungi preferred a certain water potential and temperature for sporulation. Light greatly inhibited the growth of P. lilacinus IPC-P, M. anisopliae SQZ-1-21, and L. lecanii CA-1-G but enhanced the sporulation of P. lilacinus M-14, P. chlamydosporia HSY-12-14, and L. lecanii CA-1-G.     

Tuber Blight Development in Potato Cultivars in Response to Different Genotypes of Phytophthora infestans

Migrations or introduction of new genotypes of Phytophthora infestans to a specific region imposes a different perspective for potato production. During 2009–2010, a late blight epidemic affected the Northeastern United States, which quickly spread through several states. The epidemic was characterized by the appearance of a new genotype of P. infestans designated US‐22, which was isolated from tomato and potato. Potato tubers are an essential component of late blight epidemics where the pathogen cannot overwinter on Solanaceous plants. Six potato cultivars were inoculated with 12 isolates of P. infestans (five different genotypes), including isolates of the genotype US‐22. Tuber blight development was characterized in terms of tissue darkening expressed as area under the disease progress curve values and lenticel infection. The responses indicated that US‐8 was more aggressive than US‐22, but US‐22 isolates obtained from potato were more aggressive on potato than those acquired from tomato. Tuber periderm responses to infection were limited, yet US‐8 isolates infected the periderm more often than US‐22 isolates. There were significant differences among the cultivars tested but cv. Jacqueline Lee was the most resistant overall. Although isolates of P. infestans genotype US‐22 were less aggressive in comparison with US‐8 isolates, US‐22 isolates still infected potato tubers and were as aggressive us US‐8 isolates on some cultivars. Management of late blight caused by isolates of US‐22 through host resistance may be feasible but imposes a different set of criteria for consideration from those that US‐8 imposed.     

Microsatellite markers reveal two admixed genetic groups and an ongoing displacement within the French population of the invasive plant pathogen Phytophthora infestans

Potato late blight is an example of a re‐emerging disease of plants. Phytophthora infestans was first introduced into Europe during the 19th century, where it caused the Irish potato famine. During the 20th century several additional introduction events have been suspected, especially in the mid‐70s due to the import of large quantities of potato needed after the shortage caused by drought in 1976. Here, we investigate the genetic population structure of Phytophthora infestans, at the first stages of a recent invasion process in France. A total of 220 isolates was collected from 20 commercial fields of the potato susceptible cultivar Bintje, during two consecutive years (2004 and 2005). Clustering analyses based on eight recently developed microsatellite markers reveal that French P. infestans populations are made of two differentiated genetic clusters of isolates (F_ST = 0.19). This result suggests multiple introductions of P. infestans into France, either through the introduction of a composite population of isolates or through the successive introduction of isolates having differentiated genetic backgrounds. Both clusters identified have a strong clonal structure and are similar regarding genetic diversity and mating type composition. The maintenance of differentiation between the two genetic clusters should result from the low or non‐existent contribution of sexual reproduction in French P. infestans populations.     

Concerted evolution of a tandemly arrayed family of mating-specific genes in Phytophthora analyzed through inter- and intraspecific comparisons

Multigene families are features of most eukaryotic genomes, which evolve through a variety of mechanisms. This study describes the structure, expression, and evolution of a novel family in the oomycete Phytophthora. In the heterothallic species P. infestans, M96 is expressed specifically during sexual sporogenesis, and encodes a low-complexity extracellular protein that may be a component of oospore walls. Intriguingly, M96 exists in P. infestans as 22 relatively homogeneous loci tandemly repeated at a single site, which is partitioned by inversions and retroelements into subclusters exhibiting semi-independent evolution. M96 relatives were detected in other heterothallic and homothallic oomycetes including species closely (P. mirabilis, P. phaseoli) or distantly (P. ramorum, P. sojae) related to P. infestans. Those M96 relatives also exhibit oosporogenesis-specific expression and are arrayed multigene families. Nucleotide changes and repeat expansion diversify M96 in each species, however, paralogues are more related than orthologues. Concerted evolution through gene conversion and not strong purifying selection appears to be the major contributor to intraspecific homogenization. Divergence and concerted evolution was also detected between isolates of P. infestans. The divergence of M96 proteins between P. infestans, P. ramorum, and P. sojae exceeds that of typical proteins, reflecting trends in reproductive proteins from other kingdoms. Sequence data from this article have been deposited with the GenBank Data Libraries under accessions DQ196155 to DQ196175.     

Changes of Genotype,Sensitivity and Aggressiveness in Phytophthora infestans Isolates Collected in European Countries in 1997, 2006 and 2007

A total of 241 isolates of Phytophthora infestans were collected in 1997, 2006 and 2007 in eight European countries and characterized with molecular markers (simple sequence repeats, SSR genotypes) and phenotypic traits such as sensitivity to fungicides, mating type and aggressiveness. The mating type distribution changed from mainly A1 in 1997 to a majority of A2 in 2007. No resistant isolates were detected for fluazinam and mandipropamid, whereas the proportion of isolates resistant to mefenoxam (MFX) was high and increased over the years. There was no genetic link between mating type and MFX resistance. Aggressiveness (product between lesion expansion and sporulation capacity) was slightly higher for MFX‐resistant compared to sensitive isolates and for isolates collected later compared to earlier in the same season. It was about equally high for A1 and A2 types, and for French isolates in 1997 and British isolates in 2007, but lower for French isolates in 2007. Six different SSR genotype families were distinguished. In 1997, populations were dominated by genotype families I and III/IV, which significantly declined in 2007 being largely displaced by genotype families II (‘blue 13’ type) and V, which are by coincidence mainly A2 MFX resistant and A1 MFX sensitive, respectively. However, mating type and MFX resistance were genetically not linked to SSR genotypes.     

Monitoring of Virulence in Populations of Pbytophthora infestans Using a Schwarzbach Spore Trap

A modified Sehwarzbach jet spore trap was used to sample Phytophthora infestans populations in small crops of potato within polyethylene tunnels and in the field. Airborne sporangia were eaptured in the trap and deposited upon potato leaf discs, of a cultivar lacking race-specific resistance genes, contained within the trap. Virulence characters of isolates captured in the spore trap were determined by subculturing the isolates on leaf discs of a range of potato genotypes with race-specific resistance (R-genes), incubating them and then recording compatible and mcompatible reactions, reactions. For four isolates of P. infestans the levels of sporulation on the differential leaf discs were noted and although cultivar × isolate interactions were inconsistent, variable light and temperature conditions had no significant effect on the reactions of the cultivars.     

The Influence of Temperature on Phytophthora infestans (Mont.) de Bary

The influence of temperature on the growth rate, sporulation density and zoospore release of Phytophthora infestans, cultivated on rye agar, has been studied. Temperature significantly influenced all the features of the fungus mentioned above. The highest yield of sporangia per 1 cm² of aerial mycelium occurred at 24°C while the highest percentage of sporangia releasing zoospores was observed when the fungus was grown at 15 °C. When considering the size of the fungal colony the highest production of sporangia was obtained at 20°C. It was concluded that the temperature at which the fungus was cultured predetermined the way it germinated.     

Systemically Induced Resistance in Potato Cultivars with Different Degree of Resistance to Late Blight caused by Phytophthora infestans (Mont.) de Bary

Systemically induced resistance in potato plants (Solanum tuberosum L.) against late blight Phytophthora infestans was studied macroscopically on three potato cultivars with different degrees of resistance to late blight. The cultivars, were Bintje(2), Elin(3) and Matilda(4). Figures within brackets represent resistance to leaf late blight on a scale from 1 to 5, where 5 denotes the highest level of resistance. In vitro propagated cuttings were used in the experiments, which were, performed under controlled environmental conditions. Disease resistance was induced by a local pre-infection with P. infestans or with the nonpathogen P. cryptogea. Induced resistance was observed on all cultivars. The protection causedby, induced resistance tended to be most intense on cv. Matilda, with 50–60% reduction in lesion formation 4 days after challenge inoculation.     

Seasonal change of protozoa and micrometazoa in a small pond with leaf litter supply

Protozoa and micrometazoa were observed in a small pond with leaf litter over a 2 year period. The total number of taxa observed in this pond was 83 protozoans and 30 micrometazoans, and the correspondence of each taxon and the habitats of surface water, leaf litter on the pond bottom and sedimented mud was noted. The microfauna in the litter on the pond bottom had a higher diversity than in the water or the sedimented mud. Patterns of seasonal change in the density of organisms — with one or two peaks in a year — were recognized in some taxa of protozoa and micrometazoa. Most peaks of bacterivorous protozoa in the leaf litter appeared in late autumn and spring. This phenomenon is considered to be closely related to the litter decomposition, because reduction of dissolved oxygen and/ or high bacterial density were observed in these two seasons. Nineteen species of protozoa from this pond were cultured in vitro with fallen leaves. From the results of the cultures and the observation of food vacuoles of protozoa in situ, part of the food web of the microfauna in the pond was estimated.     

Host–pathogen interaction between Phytophthora infestans and Solanum tuberosum following exposure to short and long daylight hours

The influence of short and long day length on the expression of qualitative and quantitative resistance to Phytophthora infestans in potato was studied. The incompatible interaction was tested for available set of isolates avirulent in greenhouse conditions to potato Black’s differentials possessing the genes: R2, R5, R6, R8, R9, R10, and standard potato cultivar Tarpan (no known R gene). The avirulent isolates either were completely avirulent regardless of plant growing conditions, or they infected leaflets of these differentials more frequently when plants were exposed previously to short day conditions than to long day conditions. This study highlights the importance of day length, among many other factors which are controlled, in testing the expression of the virulence of P. infestans isolates. In compatible interactions, when quantitative resistance was evaluated in differentials with gene R1, R3, R4, R7, R11, and potato cultivar Craigs Royal (no known R gene), stronger infection expressed by lesion growth rate, as well as stronger sporulation, were observed on potato leaflets of plants exposed to short day for 6–7 weeks before inoculation. The analysis of variance revealed a significant contribution to variation in lesion growth rate of day length, genotype, as well as day length by genotype interaction. Significant influence of isolate, and genotype, but not day length, on the expression of the incubation period was found. The results indicate the necessity of evaluating components of partial resistance present in potato lines used in breeding potato resistant to P. infestans in destined day length growing conditions.     

High levels of diversity and population structure in the potato late blight pathogen at the Mexico centre of origin

Globally destructive crop pathogens often emerge by migrating out of their native ranges. These pathogens are often diverse at their centre of origin and may exhibit adaptive variation in the invaded range via multiple introductions from different source populations. However, source populations are generally unidentified or poorly studied compared to invasive populations. Phytophthora infestans, the causal agent of late blight, is one of the most costly pathogens of potato and tomato worldwide. Mexico is the centre of origin and diversity of P. infestans and migration events out of Mexico have enormously impacted disease dynamics in North America and Europe. The debate over the origin of the pathogen, and population studies of P. infestans in Mexico, has focused on the Toluca Valley, whereas neighbouring regions have been little studied. We examined the population structure of P. infestans across central Mexico, including samples from Michoacán, Tlaxcala and Toluca. We found high levels of diversity consistent with sexual reproduction in Michoacán and Tlaxcala and population subdivision that was strongly associated with geographic region. We determined that population structure in central Mexico has contributed to diversity in introduced populations based on relatedness of U.S. clonal lineages to Mexican isolates from different regions. Our results suggest that P. infestans exists as a metapopulation in central Mexico, and this population structure could be contributing to the repeated re‐emergence of P. infestans in the United States and elsewhere.     

Chromosomal heteromorphism linked to the mating type locus of the oomycetePhytophthora infestans

The mating type locus of the oomycete,Phytophthora infestans, is embedded in a region of DNA that displays distorted and non-Mendelian segregation. By using DNA probes linked to the mating type locus to genetically and physically characterize that region, a large zone of chromosomal heteromorphism was detected. LocusS1 was shown to represent a tandemly repeated array of DNA that was typically present in a hemizygous state in A1 isolates while being absent from A2 isolates. The analysis of the parents and progeny of seven crosses indicated that the tandem array was linked in cis to the A1-determining allele of the mating type locus. A worldwide survey of genotypically diverse field isolates ofP. infestans indicated thatS1 was present in each of 48 isolates of the A1 mating type that were tested, but was absent in 46 of 47 A2 strains. Physical analysis ofS1 indicated that the tandemly repeated DNA sequence spanned about 300 kb and had evolved from a 1.35-kb monomer. Internal deletions occurred withinS1 during sexual propagation. This and other mutations apparently contributed to a high degree of polymorphism within theS1 array.     

A potato pathogenesis-related protein gene, StPRp27, contributes to race-nonspecific resistance against Phytophthora infestans

Late blight caused by Phytophthora infestans is the most important disease of potato. Many efforts have been made to understand molecular mechanism of the durable resistance to address the challenge raised by rapid evolution of the pathogen. A pathogenesis related protein (PR) gene StPRp27 was previously isolated from the potato leaves challenged by P. infestans. The sequence analysis and expression pattern reveal that StPRp27 may be associated with resistance to P. infestans. In present research, transient expression of StPRp27 in Nicotiana benthamiana enhanced resistance to P. infestans isolates 99189 and PY23 indicating its potential contribution to the disease resistance. These findings were also confirmed by over-expression of StPRp27 in potato cv. E-potato 3, which significantly slowed down the development of the disease after inoculation with a mixture of P. infestans races. Further, silencing of StPRp27 homologous genes in N. benthamiana harboring dominant Phytophthora resistance gene Rpi-blb1 or Rpi-blb2 showed no effects on the resistance triggered by these R genes. Our results suggest that StPRp27 contributes to a race-nonspecific resistance against P. infestans by inhibiting the disease development and has a potential use in selection and breeding for durable resistance to late blight.     

A proper PiCAT2 level is critical for sporulation,sporangium function,and pathogenicity of Phytophthora infestans

Catalase is present in prokaryotic and eukaryotic organisms and is important for the protective effects of the antioxidant system against free radicals. Many studies have confirmed that catalase is required for the growth, development, and pathogenesis of bacteria, plants, animals, and fungi. However, there has been relatively little research on the catalases in oomycetes, which form an important group of fungus-like eukaryotes that produce zoosporangia. In this study, we detected two Phytophthora infestans genes encoding catalases, but only PiCAT2 exhibited catalase activity in the sporulation stage and was highly produced during asexual reproduction and in the late infection stage. Compared with the wild-type strain, the PiCAT2-silenced P. infestans transformants were more sensitive to abiotic stress, were less pathogenic, and had a lower colony expansion rate and lower PiMPK7, PiVPS1, and PiGPG1 expression levels. In contrast, the PiCAT2-overexpressed transformants were slightly less sensitive to abiotic stress. Interestingly, increasing and decreasing PiCAT2 expression from the normal level inhibited sporulation, germination, and infectivity, and down-regulated PiCdc14 expression, but up-regulated PiSDA1 expression. These results suggest that PiCAT2 is required for P. infestans mycelial growth, asexual reproduction, abiotic stress tolerance, and pathogenicity. However, a proper PiCAT2 level is critical for the formation and normal function of sporangia. Furthermore, PiCAT2 affects P. infestans sporangial formation and function, pathogenicity, and abiotic stress tolerance by regulating the expression of cell cycle-related genes (PiCdc14 and PiSDA1) and MAPK pathway genes. Our findings provide new insights into catalase functions in eukaryotic pathogens.     

Population dynamics of Tenagomysis tasmaniae Fenton,Anisomysis mixta australis (Zimmer) and Paramesopodopsis rufa Fenton from south-eastern Tasmania (Crustacea:Mysidacea)

The population dynamics of a coastal mysid community was examined at One Tree Point in southern Tasmania over a 12-month period, including three 24-hour sampling sessions. Fourteen mysid species were recorded. Zonation patterns, life-history and diel activity of the three most abundant species, Tenagomysis tasmaniae, Anisomysis mixta australis and Paramesopodopsis rufa were examined. These three species were found in large numbers throughout the year associated with the macroalgal fringe on either side of the bay. Habitat partitioning within this zone of occurrence was evident. T. tasmaniae was the most abundant species (annual mean density 32.4 m^–3), followed by A. mixta australis (annual mean density 20.5 m^–3 ) and P. rufa (annual mean density 11.3 m^–3). The major peaks of abundance of these three species were temporally separate. Breeding was intensive for the three species from spring till late autumn. A cessation of breeding in winter was evident for A. mixta australis and to a lesser extent for P. rufa but T. tasmaniae bred throughout the year although at a reduced rate during winter. Examination of the diel behaviour identified peaks of abundance at sunrise and sunset for T. tasmaniae; P. rufa was caught in greater numbers at night, but no consistent pattern was observed for A. mixta australis.