الفهرس 
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Abstract
An isolate of Bacillus megaterium efficient in dissolving phosphate was obtained from the Microbial collection of Department of Agric. Microbiology, Fac. Agric. Minia University.
Bacteriophages specific to B. megaterium were enriched from a soil sample collected from the rhizosphere of maize plants, growing in the Experimental Farm of the Faculty of Agriculture, Minia University,.
The spot test was used for detection of phages. Bacteriophages of B. megaterium were found to be of widespread occurrence in soils from where the samples had been taken.The single plaque isolation technique was used to purify phages. Seven single plaques of phages specific to B. megaterium each having different morphology were picked and kept as single phage isolates.
Five hundred ml of high titre phage suspension were prepared for each of the seven phage isolates. The titres of the prepared phage suspensions ranged from 3.4x1010 pfu/ml to 6.4x1012 pfu/ml.
The different characteristics (optimum pH, thermal inactivation point, sensitivity to ultraviolet irradiation, host range and particle size and morphology) of the 7 phage isolates were studied to find out if these phages are different types or similar.
On the basis of the optimum pH values of each phage isolate the seven bacteriophage isolates were divided into four groups (group A, B, C and D). Each group contained the phage isolates which exhibited the same optimum pH.
The thermal inactivation point of each phage isolate was studied. According to the similarity in the thermal inactivation point, bacteriophages were grouped. The seven bacteriophage isolates were divided into four groups (group A, B, C and D). Each group contained the phage isolates which exhibited the same thermal inactivation point.
Sensitivity of the isolated phages to UV radiation (at wave length of 254 nm) was studied. UV inactivated the isolated phages at different exposure time. Accordingly, the isolated phages were divided to four groups (group A, B, C and D). Each group comprised the phage isolates which inactivated after the same exposure time.
Interestingly, the four groups of phages which divided on the basis of the optimum pH, thermal inactivation point were found to be the same as those classified on the basis of the sensitivity to U.V. radiation. Such results may indicate that the phages of each group are belonging to a single phage type.
Bacteriophage isolates specific to B. megaterium were negatively stained and examined by electron microscope. All phage isolates were found to be of head and tail type. Phages of each group (divided on the basis of the thermal inactivation point, sensitivity to UV radiation and optimum pH) were found to be morphologically similar. The differences in head and tail dimensions of the phages of each group are within the standard deviation and are not statistically significant. Therefore, on the basis of the above mentioned information, it can be concluded that the phages of each group represent one phage type. i.e. the seven isolated phages of B. megaterium were found to be belonging to four phage types. The four phage types of B. megaterium were designated Bm1, Bm2, Bm3 and Bm4 for phages of group A, B, C and D, respectively.
Each of the four phage types of B. megaterium was tested against four different bacillus species. all phage types were infectious to B. megaterium (the main host). Whereas, none of the four phages was infectious to the other tested bacillus species with exception of phage Bm3which was infectious to B. ceureus.
In an attempt to protect Bacillus megaterium against phage infection, Bacillus megaterium was prepared in form of alginate immobilized cells. In addition, a phage resistant mutant of Bacillus megaterium was successfully isolated.
The efficiencies of Bacillus megaterium in the two forms of inocula (immobilized cells and the phage resistant mutant) in dissolving phosphate and their susceptibility to bacteriophages were tested in pure liquid culture (in vitro) and under cultivated soil condition (in vivo).
The presence of phages had no effect on the efficiency of the phage resistant mutant and the immobilized cells in dissolving insoluble phosphate.
Since the immobilized cells of Bacillus megaterium and its isolated phage resistant mutant exhibited high resistance to the virulent phages and high biological activities under aseptic conditions, it was of a particular interest to evaluate their efficiency under cultivated soil conditions.
In soil cultivated with maize plants, densities of sporeforming bacteria in rhizosphere soils of the plants inoculated with the different forms of inocula were tested.
The obtained results indicated that at any sampling time, rhizosphere of maize plants inoculated with the immobilized cells contained much higher numbers of sporeforming bacteria than in those inoculated with the free cells.
Presence of phages, markedly reduced numbers of sporeforming bacteria in rhizosphere soils of maize plants inoculated with B. megaterium in free states, as compared to those inoculated with free cells in absence of phages. Furthermore, in plants inoculated with immobilized cells of B. megaterium plus their specific phages, slight reduction in numbers of sporeforming bacteria in rhizosphere soils was observed, as compared to those inoculated with immobilized cells in absence of phages.
No pronounced differences in numbers of the sporeforming bacteria were detected in rhizosphere soil of maize plants inoculated with the phage resistant mutant plus phages as compared to those inoculated with these mutant in absence of phages.
A marked increase in numbers of sporeforming bacteria in rhizosphere soils of maize inoculated with the immobilized cells of B. megaterium was observed as compared to those inoculated with the phage resistant mutant either in presence or in absence of phages.
At any sampling time, the rhizosphere soil of maize plants which inoculated with the immobilized cells of B. megaterium contained much higher numbers of sporeforming bacteria than those inoculated with the free cells.
Moreover, in absence of phages inoculation of maize plants with the wild types (free cells) or phage resistant mutant of B. megaterium led to significant increase in P- content, fresh and dry weight of the plants as compared to the uninoculated ones.
In addition, at any sampling time the P-contents, fresh and dry weight of maize plants inoculated with the wild type plus phages was lower than in the other treatments.
On the basis of the obtained results it can be concluded that, the depressive effect of the bacteriophages can be avoided by application of the bacterial inocula in form of alginate immobilized cells. Moreover, isolation of phage resistant mutants of such desired bacteria can be used as well to avoid the phage attack.
Therefore, application of immobilized cells or phage resistant mutant of such desired bacteria as biofertilizers is highly recommended to avoid the phage attack and to promote the efficiencies and maintenance of these microorganisms in the soil.