Mating Types, Race and Genetic Variability in Phytophthora Infestans, the Cause of Late blight of Potato

Abstract

Late blight, caused by the fungus, Phytophthora infestans is a significant threat to potato, which is an important vegetable crop of Pakistan and has assumed strategic significance from food security point of view due to its high-energy yield per unit area of production. P. infestans causes severe defoliation and also infect the potato tubers, when the inoculum moves through the soil via water channels. The disease can destroy the whole crop within a week or two. The high degree of the destructiveness of this disease is mainly due to the heterothallic nature of the fungus and occurrence of sexual reproduction in its population. Oospore produced as a result of pairing of opposite mating types (A1 and A2) are tough and hard survival structures and are resistant to fungicides used for its control. The new population produced as a result of sexual reproduction is mostly fungicide (metalaxyl) resistant and is more diverse genetically. Genetic diversity in the population mainly affects the virulence pattern of the fungus and new population is mostly more aggressive than the older one. Resistant and diverse inoculum is the main reason of the failure of efforts to manage the disease. Studies in Pakistan have shown the presence of both A1 and A2 mating types. This necessitated the investigation of temporal and geographical distribution of the mating types and analysis of the resistance of P. infestans to widely used fungicides metalaxyl for developing appropriate management strategies, especially based on integrated approach with primary focus on genetic manipulation and fungicide resistance management. It is with this objective that current project was conceived. To collect the samples of late blight, areas/locations of Punjab, lower valleys of North Western Frontier Province (NWFP), Azad Jammu and Kashmir (AJK), Kaghan valley, central and upper Swat valleys were surveyed during the year 1997-1998. During the year 1999-2000, areas of Punjab central and upper Swat valleys were surveyed. From these samples, 281 isolates were obtained. Eighty isolates were obtained from the samples collected during 1997-1998, 161 isolates were obtained from the samples collected during 198-1999 and 40 isolates were obtained from the samples collected during 1999-2000. All were characterized on the basis of mating types and metalaxyl resistance. Race pattern of 140 isolates and protein profiles of 49 isolates were determined. Both mating type of P. infestans were found in all the areas of country, except in lower valleys of NWFP. During the year 1997-1998, frequently of A2 mating type was highest in the population of AJK, i.e., 56.2%. In Punjab and Kaghan valley, the frequently of A2 was lower i.e., 35% and 42% respectively. The pattern of this distribution changed during the next year (1998-99). In Punjab, the frequently of A2 slightly increased (38%), whereas in Kaghan valley it was considerably increased (61%). In Azad Jammu and Kashmir (AJK) frequently of A2 decreased to 41%. During the year 1999-2000, self-fertile isolates were also found in addition to two mating types, in the isolate and self-fertile (30%). Three phenotypes of the fungus (with respect to its response to metalaxyl i.e., sensitive, intermediate and resistance) were observed among the isolates of almost all the areas. The intermediates were of higher frequency as compared to other two phenotypes among the isolates of all the areas during the whole period of study (from 1997-2000). Frequency of sensitive isolates decreased (from 16.25% to 2.5%) throughout the years of study. Frequency of resistant isolates decreased by 9% in one year (from 1997-1998 to 1998-1999), whereas increased by 19% in one year (from 1999-2000). During the year 1997-1998, highest frequency of resistant isolates was observed in the population of Punjab i.e., 17% whereas lower frequencies were observed in the population of AJK and Kaghan valley i.e., 6.2% and 8.3% respectively. However, this pattern also changed during the next year (1998-1999). In Punjab, frequency of resistant isolates decreased to 4% whereas Kaghan valley it increased up to 10.7%. In Punjab, where the frequency of A2 was somehow stable during two years, frequency of resistant isolates decreased, which might be due to increased frequency of intermediates (92%) produced as a result of sexual reproduction between sensitive and resistant isolates. In Kaghan valley, frequency of resistant isolates and frequency of A2 mating type increased simultaneously. As A2 is reported to be more aggressive and resistant, so increase in its frequency may be responsible for increased resistance in this area. Frequency of resistant A2 (RA2) WAS ALSO HIGHER (8.33% in 1997-98 and 10.7% in 1998-99) in this area as compared to others during both years. During the year 1999-2000, only the population of Punjab was characterized. Frequency of sensitive isolates decreased from 4% to 2.5%, whereas frequency of resistant isolates increased from 4% to 22.5% during this year, which indicated the gradual establishment of resistant strain. Highest proportion of intermediate response was observed among the isolates of both mating types from all the areas, which is also believed to be the result of sexual reproduction between sensitive and resistant isolates. Continuous monitoring of mating types of P. infestans and its resistance to metalaxyl is necessary to gain the knowledge about the current situation of resistance and for developing new disease management approaches and resistance management. Thirty-nine races were identified in a population of 140 isolates of the fungus, indicating a high degree of variability. Complex races were more common, showing high level of aggressiveness in population of P. infestans. Occurrence of different races in all the areas may be due to occurrence of sexual reproduction and import of different strains through international seed. Few races were also common in different areas, indicating the transfer of races from one area to another mainly through seed flow. In the isolate collection of year 1999-2000 diversity was less as compared to previous two years collection, but complexity increased i.e., more complex races were of highest frequency. Isolates of Phytophthora infestans from Pakistan and Cornell University, USA were used for SDS-PAGE analysis, Polyacrylamide gel (15%) was used to compare the protein profile of these isolates. Ten band were found polymorphic and scored to generate dendrogram. Two majors’ groups (A and B) were observed with help of cluster analysis. Group A further divided into six and group B intro three sub-group. All isolates from Cornell University and 50% isolates from Pakistan were grouped in the same cluster. This indicating a close relationship of Pakistani population with that of USA in the same cluster indicating high level of homology among each other. Variability in the DNA profile of Phytophthora infestans isolates is being further studied.