الفهرس 
INTRODUCTION
The importance of potato in EgyptOnly 14 pages are availabe for public view
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Abstract
Bacterial soft rot disease, caused by Erwinia carotovora subsp. carotovora and Erwinia carotovora subsp. atroseptica are the most important and widespread bacterial disease to a wide variety of plants either in the field or transit or during storage causing great losses in potato tubers.
Results of this work can be summarized in the following:
1. Fifty bacterial isolates were obtained from naturally diseased potato tubers and plants, cabbage, carrot, and lettuce showing symptoms of soft rot and blackleg disease collected from different localities of El-Minia, Assiut and Sohag Governorates during 2010-2011 seasons. Twenty eight isolates were proved to be pathogenic and produced symptoms of soft rot on potato tubers. They varied in their virulence and identified as E. carotovora according to their morphological, physiological, and biochemical characteristics.
2. Ten arbitrary oligonucleotide primers were used in the present investigation to generate RAPD profiles from six isolates of Erwinia carotovora. Only three primers [(1): CUGEA-4, (2): CUGEA-5, and (3): CUGEA-6] were amplified successfully on the genomic DNA from taken isolates yielding distinct RAPD patterns. The number of the amplified fragments per primer varied between two and eight, with an average 22 bands per primer. These fragments have a size ranged about from 1156 to 218 bp. A total number of 67 bands were amplified with no band being monomorphic and all bands were polymorphic with a polymorphism ratio of 100%.
Primer 2 (CUGEA-5) generated the largest number of fragments (24), Primer 3 (CUGEA-6) produced (23) fragments, while primer 1 (CUGEA-4) generated the lowest number of fragments (20). The three primer produced a total number of 98 band in the six isolates of Erwinia carotovora isolates.
a- The first Primer (1): CUGEA-4
Primer (1): CUGEA-4 produced 20 bands in the six isolates ranged in size about from 1108 to 376 bp. The number of fragments generated by this primer varied among isolates where the lowest no. was three which observed in isolate no. 6, while the highest no. was eight which observed in isolate no. four. This primer could be used as genetic marker to distinguish Ec. subsp. carotovora by generates bands with size about 412, 431 and 542 bp which generated only with subsp. carotovora. Moreover, these bands could be used to distinguish isolates of Ec. subsp. carotovora according to their virulence, where bands with size 412 and 431 bp only generated with the high and intermediate virulent isolates, respectively and band with size 542 bp produced only with low virulent isolates.
b- The second primer (2): CUGEA-5
Primer (2): CUGEA-5 produced thirty bands in the six isolates having size about of 1156 and 247 bp. The number of fragments generated by this primer varied among isolates from 2 in isolate nos. 5 to 8 in isolate no. 1.
c- The third primer (3): CUGEA-6
Primer (3): CUGEA-6 produced 39 bands in the six isolates ranged in size about from 1068 to 218 pb. The number of fragments generated by this primer varied among isolates where the lowest no. was 5 which observed in isolate no. 3, while the highest no. was 8 which observed in isolate no. 2. The second and the third primers could be used as genetic markers to distinguish the isolates of Ec. subsp. atroseptica according to their virulence, where primer no. 3 generated band with size 964 bp with high virulent isolates. While primer no. 2 generated band with the same size (962 bp) which sign to the low virulent isolates.
d- All isolates divided into two main clusters, according to genetic relationship showed in the dendrogram. The first cluster contains isolate no. 1 which represents the main root of dendogram. Subsequence by the second cluster of isolates no. 2 with 20% of genetic similarity. Both isolates no.1 and 2 are highly virulence. The other isolates divided into two sub-clusters at 25% of genetic similarity. The first sub-cluster composed isolate no. 4 and the second sub-cluster divided into two groups at 40% genetic similarity. The first group contains isolate number 3 which represents the ancestor of the second group. The second group contains two isolates (nos. 5 and 6) which represents the most recent isolates in the dendogram with 70% genetic similarity, both isolates are low virulence.
3. The amplification results of the 16s rRNA gene of the six Erwinia
carotovora isolates, as obtained by specific primers shown that insertion
or deletion were not encountered. The phylogeny tree showed harmony distribution of the isolates with the sequences of Erwinia sp. obtained from GenBank. Moreover, new evolutionary taxonomy of Erwinia could be shown.
4. Nine isolates of Erwinia carotovora were used in the investigation of protein assay. Protein of purification fractions separated by SDS-PAGE analysis of the molecular weight in Kda showed that the number of protein fractions per isolate varied between 11 to 18 with an average of 14 fractions per isolate. These polypeptide fractions have a size ranged from about 185 to 56 Kda.
A total number of 129 fractions were shown in the nine isolates with 36 fractions being monomorphic fractions, and the other 93 fractions were polymorphic. Some isolates could be distinguish by some polypeptide fractions as instances, fractions with about 172, 176 and 170 Kda were found to be present only in isolates 6, 7 and 9, respectively and could be used to distinguish isolates collected from Sohag Governorate. Also, fractions with about 150 Kda found only in isolates 1 (Ecc4) and 4 (Eca25) could be used to distinguish the high virulence. Also, fractions with about 150 Kda present only in isolates 1 and 4 could be used to distinguish the high virulence. Also, the polypeptide fractions with about 78 and 79 Kda were present only in isolates 5 (Eca35), 9 (Eca26) and 8 (Eca42) and could be used to distinguish isolates E. carotovora subsp. atroseptica. Moreover, polypeptide fraction with about 56 Kda is distinguish only isolate 8.
Results also indicated that polypeptide fraction with about 165 Kda was absent in isolates 4, 6 and 9 but present in the others that could be used to distinguish isolates of Eca. Also, polypeptide fractions with about (130, 155), 145, (107, 70), 88 and 68 Kda were absent in isolates 3 (Ecc11); 6 (Eca40); 1 (Ecc4); {5 (Eca35), 6 (Eca40)} and {1 (Ecc4), 6 (Eca40)}, respectively.
a- A phylogeny tree, indicating genetic relationships between the tested Erwinia carotovora isolates. Isolates were classified into two groups. The first group containing isolates no. 1, which represents the main root of this cluster, and isolates nos. 6, 7, 2 and 3. The second group containing isolates nos. 5, 8, 4 and 9. This group divided into two sub-clusters, one of them containing isolates nos. 5 and 8 and the other containing isolates nos. 4 and 9.
5. The severity soft rot disease was significantly lower in tubers wounding after soaking than wounding before soaking.
6. Transmission electron micrography showed that silver nanoparticles are nanosized and well dispersed. The size of the silver nanoparticles were examined and it was found to be of: 26.1, 29.6 and 32.1 nm in size.
7. All tested concentrations of silver nanoparticles exhibited different degree of bacterial growth. The growth inhibited as the concentration increased. The concentration of 100, 150 and 200 µg ml-1 caused complete inhibition for growth of both tested isolates.
8. The minimum bactericidal concentration (MBC) of silver nanoparticles for suppression growth of Ecc6 and Eca19 were 100, 150 and 200 µg ml-1. They completely inhibited growth of tested isolates. The other tested concentrations (50, 60, 80, and 90 µg ml-1) showed partial effect on inhibiting colony growth. 50 µg ml-1 of silver nanoparticles caused the highest number of colony numbers followed by 60, 70, and 80 µg ml-1. 90 µg ml-1 showed the least number of colonies of tested isolates.
9. TEM analysis clearly showed that the nanoparticles were accumulated in the bacterial cell membrane, while some of them successfully penetrated into the cells. In addition, leaking of intracellular substances and coagulation of nanosized particles at the bacterial surface are also seen in the same TEM micrograph.
10. Soaking potato tubers for 15, 30, and 60 min. at conc. of 200 µg ml-1 of silver nanoparticles caused completely suppression tubers soft rot for all tested storage periods. Also, soaking potato tubers for 60 min. at conc. of 150 µg ml-1 showed complete suppression of the disease. 100 µg ml-1 for all tested soaking exhibited partial suppression of soft rot disease for all tested storage periods.
11. The effect of Degaclean on growth of Ecc6 and Eca19 at concentrations of 0.3%, 0.6%, 0.9%, 1.2%, 1.5% and 2% was tested in vitro. Degaclean at conc. of 2% completely killed all bacterial cells. Data also showed that the other tested concentration of Degaclean exhibited partial inhibition of Ecc6 and Eca19 growth. Also the increase of Degaclean concentration was correlated with decreasing bacterial growth. Conc. of 1.5% and 1.2% of Degaclean caused the highest growth inhibition followed by 0.9%, 0.6% conc. and finally 0.3% conc.
12. Soaking potato tubers in 0.5, 1.0 and 2.0% Degaclean for 30 min have significantly reduced soft rot development in potato tubers. 2% Degaclean completely suppression tubers soft rot for all tested storage periods. Data also revealed that 1.0% Degaclean caused highest decreased in disease severity followed by conc. of 0.5% Degaclean. All treated potato tubers soaked in conc. of 0.5, 1.0 and 2.0% Degaclean, and stored for 0 month showed the highest decreased in disease severity followed by those stored for 1 and 2 months as compared with those of 3 month storage period.