الفهرس 
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Abstract
Pythium is a large and important genus in the class Oomycetes and its species are cosmopolitan and can be found as saprophytes, parasites, facultative parasites or mutualistic; aquatic, amphibious or terrestrial fungi. They occur most abundantly in cultivated soils near the root region and less commonly in non-cultivated soils. Most of Pythium species were characterized by their pathogenicity to many plants. Whereas some others species such P. oligandrum are characterized by their ability to parasitize other pathogenic fungal species. Although there are previous studies on the isolation and identification of Pythium species in Egypt, there are no sufficient physiological studies on the pathogenic and the mycoparasitic species to provide information on the differences between them which is important for efficient use of the mycoparasitie in biological control. This work was undertaken to provide such information as well as confirm the identification of Pythium species isolated using molecular technique. Pythium species were isolated from rhizosphere soil of 16 plant species cultivated in nine provinces of El- Minia governorate during 2006-2008. These plant species were Abelmoschus esculentus, Allium cepa, Brassica oleracea, Cucumis sativus, Eruca sativa, Glycine max, Lactuca sativa, Phaseolus vulgaris, Raphanus sativus, Sacharum spontanum, Sorghum bicolor, Spinacia oleracea, Trifolium alexandrinum, Triticum aestivum, Vicia faba L. and Zea mays.
Results indicated that 746 isolates belonging to 11 species and three groups of Pythium were isolated using selective agar medium (NARM) by three technique of isolation, which are; zoospores baiting technique (ZBT), direct isolation from soil (DIS) and serial dilution method (SDM). Isolated species were identified primarily on the basis of morphological characteristics and were confirmed by molecular identification by sequencing of rDNA-ITS including the 5.8 S rDNA of Pythium species. Eleven Pythium spp. were identified which were designated as (MS14, MS09, MS27, MS26, MS42, MS43, MS36, MS31, MS87 and MS133) and the sequencing data showed complete similarity (100%) to the known reference Pythium species in Gen-Bank data, according to accession numbers of AB274404.1 as P. aphanidermatum, AF597536.1 as P. aquatile, AY597756.1 as P. catenulatum, AY598689.1 as P. deliense,. AB597526.1 as P. graminicola, AF452142.1 as P. irregulare, AB989556.1 as P. myriotylum, AY986954.1 as P. oligandrum, AY598701.1 as P. spinosum var. spinosum, and AY598657.1 as P. ultimum var. ultimum. Only the isolate MS51 showed 99% similarity to Gen-Bank accession number, AY598690.1 as P. diclinum. While Pythium “group F”, Pythium “group G” and Pythium “group P” were only morphologically identified.
Occurrence of Pythium species (number of isolates obtained) in different localities in El-Minia governorate revealed that; P. aphanidermatum was generally the most dominant species with 142 isolates representing 19.03 % of total isolates, followed by P. ultimum var. ultimum (117 isolates representing 15.68% of total isolates of Pythia). P. diclinum was the third species with 88 isolates representing 11.79%. P. graminicola and P. myriotylum were the lowest frequent species with occurrence percentage (1.47% and 2.01%) of total isolations, respectively.
P. aphanidermatum was the predominant species in most provinces of El- Minia governorate with high frequency reaching to 27.5% of Pythium isolates in El- Minia city. On the other hand, the highest frequency isolation of P. ultimum var. ultimum was in Maghagha and Beni-Mazar with isolation percentage of 26.39 and 21.52%, respectively. The highest isolation frequency for P. diclinum, P. deliense and P. oligandrum were in Beni-Mazar representing 21.52, 12.66 and 11.39%, respectively, while the highest isolation frequency of P. irregulare and P. spinosum var. spinosum were recorded in Abou-Qurqas representing 16.1 and 14.8%, respectively.
Occurrence of Pythium species in correlation with plants rhizosphere soil revealed that P. aphanidermatum was the only predominant species in rhizosphere soil of Glycine max with occurrence percentage of 64.71%. Moreover, P. deliense, P. diclinum and P. ultimum var. ultimum were more frequent in rhizosphere soil of Zea mays, Cucumis sativus and Phaseolus vulgaris with occurrence percentage of 24.39, 20.69 and 38.78%, respectively. On the other hand, the highest frequencies in case of P. oligandrum and Pythium “group P” were recorded in rhizosphere soil of Eruca sativa and Raphanus sativus with occurrence percentage of 34.15 and 18.52%. While P. irregulare and P. spinosum var. spinosum were the most frequent species in rhizosphere soil of T. alexandrinum with occurrence percentage of 15.87 and 14.29%, respectively. Other Pythium species like P. aquatile, P. catenulatum, P. myriotylum and Pythium “group G” were the most frequent species in rhizosphere soil of S. spontanum plants with occurrence percentage of 16.33, 14.29, 10.20 and 12.24%, respectively.
The seasonal fluctuation of Pythium species showed that P. ultimum var. ultimum, P. spinosum var. spinosum, P. irregulare, P. catenulatum, and P. aquatile were mostly frequent in winter, while P. oligandrum, Pythium ”group P” and Pythium ”group F” were the most abundant in spring. On the other hand, P. aphanidermatum and P. deliense were the dominant species in summer. In general, Pythium spp. were most dominant in winter but showed the lowest occurrence in summer.
Investigation on the effect of temperature on the isolated Pythia showed that, P. aphanidermatum and P. deliense had higher temperature maxima (43°C) than the other Pythia studied. The optimum temperature for all the species lied between 25°C and 30°C, while the minimum temperature supporting mycelial growth was around 5°C expect for P. aphanidermatum which did not grow below 10°C.
For biological control of pathogenic Pythia species, three isolates of P. oligandrum (MS15, MS19 and MS31) were used. These isolates were selected as the strongest antagonists to phytopathogenic Pythia to be used in the pathogenicity, physiological and antagonistic experiments against the plant pathogenic P. ultimum var. ultimum (MS133) and P. aphanidermatum (MS14).