الفهرس 
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Abstract
In this study A bacterial isolate was picked from sewage water sample collected from sewage treatment plant of El-kawther City, Sohag, Egypt. A smear of the isolated bacterium was prepared on a glass slide and Gram stained. This isolate was found to be a rod-shaped Gram-negative aerobic bacterium. The isolated bacterium was identified in and was found to be Pseudomonas aeruginosa (P. aeruginosa).
Sensitivity of P. aeruginosa to 19 antibiotics was tested. Among the 19 antibiotics tested, 2 antibiotics exhibited inhibitory effect against P. aeruginosa. Ciprofloxacin was found to be the most efficient antibiotic against P. aeruginosa, followed by Meropenem.
P. aeruginosa was found to be tolerant to salinity stress. At any of the tested concentrations of NaCl (1 – 9%) the growth of P. aeruginosa tended to increase with increasing the incubation period. The highest growth at any NaCl concentration was recorded at the 6th day of incubation at 37ºC. Moreover, among the different NaCl concentrations tested, the highest growth of P. aeruginosa was recorded at 1% NaCl.
P. aeruginosa was found to be tolerant to all tested pH levels (pH 4 – 10). At any pH level the growth tended to increase with increasing the incubation period. The highest growth value was recorded at pH 6 after incubation for 6 days.
Bacteriophages specific to P. aeruginosa were successfully isolated from the sewage water sample collected from sewage treatment plant of El-kawther City, Sohag, Egypt. Phages of P. aeruginosa were found to be common in the collected sample. The single plaque isolation technique was used to obtain pure phage isolates of P. aeruginosa. Twenty single plaques morphologically different were selected and kept as pure phage isolates. The isolated phages formed circular single plaques of 1 to 3 mm in diameter and clear in appearance.
One hundred ml of high titer phage suspension were prepared for each phage isolate of P. aeruginosa. The titers of the prepared suspensions of the twenty phage isolates specific to P. aeruginosa were ranged from 43 x108 to 58 x108 pfu/ml.
The different characteristics of the twenty phage isolates were studied to find out if these phages are different types or similar.
The infectivity of the twenty phage isolates was studied at various pH levels (pH 4 – 12). At any pH level tested, all phage isolates formed lysed spots. i.e. the isolated bacteriophages of P. aeruginosa are tolerant to alkaline and acidic reactions. The twenty phage isolates were classified into five groups (A, B, C, D and E) each group comprised number of phage isolates of the same optimum pH for infection. Since the optimum pH for phage isolates of each group varied from group to another, it is likely for the isolated phages to be belonging to five different phage types, i.e. the phages in each group represent one phage type.
The thermal inactivation point of each phage isolate was estimated. According to the similarity in the thermal inactivation point, phages were divided to five groups. Groups (A, B, C, D and E) each comprised phage isolates of the same thermal inactivation point. Interestingly, the five groups of phage isolates which divided on the basis of the optimum pH were found to be the same as those classified on the basis of the thermal inactivation point. These results may indicate that the twenty phage isolates of P. aeruginosa are belonging to five phage types. i.e. phage isolates in each group represent one phage type.
Sensitivity of the isolated phages of P. aeruginosa to UV radiation (at wave length of 254 nm) was studied. The U.V. radiation at wavelength of 254 nm inactivated the isolated phages at different exposure time. Accordingly, the isolated phages of P. aeruginosa under study were divided to five groups. Each group comprised the phage isolates which inactivated after the same exposure time.
The five groups of phage isolates which divided on the basis of the optimum pH and 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 P. aeruginosa were negatively stained and examined by electron microscope. All phage isolates were found to be of head and tail type. Phages of each group were found to be similar in their dimensions. Interestingly, phages of each group (divided on the basis of the optimum pH, thermal inactivation point and sensitivity to UV radiation) were found to be morphologically similar. Therefore, on the basis of the obtained results it can be concluded that the twenty phage isolates of P. aeruginosa represent five different phage types. These phage types of groups A, B, C, D and E were designated Pa 1, Pa 2, Pa 3, Pa 4 and Pa 5, respectively.
Phages as antimicrobial agent:
An experiment was carried out as an attempt to eliminate P. aeruginosa contaminating milk and water.
The initial count of P. aeruginosa in contaminated milk and water was 35x103. In absence of the specific phages gradual increase in number of P. aeruginosa was recorded with the laps of time in case of incubation at 37 and 7ºC. The highest numbers were recorded in both water and milk after 24 h.
On the other hand, with application of phages to the contaminated milk and water, gradual reduction in numbers of P. aeruginosa was recorded. After 24h. no bacterial cells were detected neither in milk nor water incubated at 37 or 7ºC.
Moreover, in contaminated milk and water which treated with the specific phages, numbers of phage particles were estimated at zero time and after incubation for 24h at 37 and 7ºC. Number of phage particles at zero time was estimated to be 29x107 pfu/ml. Whereas, high numbers of phage particles were detected in both milk and water by the end of the experiment (after 24 h).
Generally, it can be concluded that P. aeruginosa was found to be of wide spread occurrence in sewage water. This microorganism is known to be resistant to most antibiotics, highly tolerant to wide range of pH levels and salinity stress. The use of bacteriophages specific to P. aeruginosa as antimicrobial agent was found to be effective to get rid of such pathogenic bacterium in contaminated water and milk. Therefore, application of specific phages to foodstuffs is highly recommended to avoid contamination or to get rid of this pathogenic bacterium.