Among the 7 wastewater indigenous isolates only two (Rz6 and Rz7) as well as two exogenous strains (S1 and PS) showed the ability to clarify domestic wastewater indicating that they possess the required degrading enzymes. Molecular characterization of the 4 most active bacteria revealed their belonging to Escherichia coli
(Rz6), Providencia vermicola
sp. (S1) and Pseudomonas
sp. (PS). The mixed culture (combination with a four selected cultures) proved to be the most efficient for decontamination of domestic wastewater in the present study. The marvellous resistance and superior potentiality of the present bacterial selection for biodegradation of toxic organic and uptake of inorganic pollutants were reported earlier [9
spp. such as Rz5 and S1 are well known as highly resistant spore-forming bacteria that possess excellent characteristics and extremely efficient for many agricultural, environmental and industrial applications [20
]. Results confirmed the great and remarkable ability of the selected Bacillus sp. to degrade all the investigated contaminants at very fast rates which explains their occurrence in the highly contaminated and hypertrophic environments such as Lake Mariut, a brackish water lake in south Alexandria, Egypt.
(PS) is a Gram-negative, rod-shaped, motile, single polar-flagellated, soil denitrifying bacterium with superior biodegradation and transformation ability for many environmental pollutants [27
]. Active and dead masses of Pseudomonas spp. are well known as biodegraders for pesticides [10
], crude [9
] and vegetable oil [12
Wastewater treated in the present study can be classified as moderately strong depends on levels of contaminants it contains that required powerful treatment to minimize its pollution load and discharge it safely. Batch treatment using free living individual and mixed selected bacteria aimed to design an efficient treatment process for eliminating or minimizing chemical, biological and organic load from the drainage network in Jeddah City to protect the receiving ecosystem. It was time and species dependent and subsequently resulted in varied removal efficiencies of contaminants.
Total suspended solids (TSS), biological oxygen demand (BOD) and chemical oxygen demand (COD) are three major waste characteristics determining the pollution levels in wastewater. They are also the main identification indicators for determining the efficiency of any proposed treatment system. The maximum permissible limits (MPLs) of these indicators are specified nationally by laws and regulations as those of Egypt and Saudi Arabia to provide a safe discharge. PS considered the most efficient for removing organic matter (BOD and COD) brought them to safe discharge limits in the batch treatment. However, S1, Rz6, Rz7 and the mixed culture showed the highest removal of TSS, FOG, TC and FC respectively but their residuals still above their MPLs for the safe discharge (23.3, 20 and 200 fold for TSS, FOG and TC respectively).
Microbial degradation requires optimum pH and temperature to proceed. In the present study pH range (5.60 to 6.90) and optimum temperature of 37°C encourage all the selected bacteria to remove organic matter and other included pollutant which is supported by other workers [28
]. However, pH of the treated domestic effluent lies in the permissible limits (6-9) stated by the Presidency Meteorology and Environment Organization (PME), KSA. Environmental laws stated that DO content of water and wastewater should not decrease below 5 mg/l to be safely discharged. Raw wastewater recorded 3.5 mg/l DO that is lower than DO MPL (5 mg/l). More oxygen is required for degradation of the included organic matter to the extent that wastewater may be depleted of oxygen during stabilization. Therefore, high amounts of oxygen in addition to powerful bacterial strains are needed for degradation of the contaminants. Total suspended solids TSS include settle able and colloidal fractions. The colloidal fraction cannot be removed by settling. Generally, biological oxidation or coagulation, followed by sedimentation, is required to remove these particles from suspension. The highest TSS RE recorded was 39.1% achieved using the free living Bacillus amyloliquefaciens
(S1) after 7 day which is 23.3 fold higher than its maximum permissible limit (MPL) of 60.
(PS) achieved the highest RE of BOD (87.5%) after 7 days produced an effluent with good quality (55 mg BOD/l) that can safely be discharged into open systems. BOD is removed mainly by the powerful ability of the augmented bacteria that metabolize organic matter found in wastewater transforming them into harmless by-products such as carbon dioxide and water. During the first 24 h, BOD of wastewater augmented with B. amyloliquefaciens
, P. stutzeri
and the mixed culture as well as the control increased at varies levels (9.5, 0.4, 0.4 and 11.3% respectively). The control continued to have higher BOD reaching the highest increase (-148.7%) after 7 exposure days. These increases reflect toxicity of the wastewater that led to the death of some bacterial cells with the addition of their organic matter to the wastewater as BOD. It is also reported that unless the microbial cells produced during organic matter decomposition are removed from the solution, complete treatment will not be accomplished because biomass of these cells will be measured as BOD in the effluent. In such case the only treatment that has been achieved is that associated with the bacterial conversion of a portion of the organic matter originally present to various gaseous ends by products [29
Raw wastewater recorded relatively high COD level (925 mg/l) at the zero time that was regularly decreased till the 7th
exposure day reaching 65 mg/l (93.0% RE) achieved by Pseudomonas stutzeri
(PS). The efficiency of P. stutzeri
(PS) in removing COD load is also supported by Ramteke et al. [30
] for heavily polluted tannery effluents. According to the MPL of the COD (100 mg/l in the Egyptian regulations) the lowest recorded RC (65 mg/l) of the COD is much lower and compiles with the law for safe discharging into open environments.
Raw wastewater recorded very high FOG level (2000 mg/l) at the zero time that was regularly decreased till the 7th
exposure day reaching 200 mg/l (90.0% RE) achieved by Escherichia coli (RZ6) indicating high capability of the selected bacteria to use fatty organic matter as a source of carbon and energy [11
]. The lowest FOG residue recorded 200 mg/l which is 20 fold higher than the MPL of FOG in the final treated effluent (10 mg/l Egyptian Limits) and 1.7 fold higher than Saudi Limit (120 mg/l) stated for the discharge into Central Treatment Unit [31
Toxicity of wastewater on the selected bacteria was determined as growth inhibition (GI) shown in the total viable count of bacteria (TVC). GI gradually increased reaching the highest GI after 7th
day. In that respect, mixed culture recorded the highest (99.1%) growth inhibition after the 7th
day. Control wastewater showed relatively lower GI (75% after 7 days) compared to the seeded wastewater. These results are attributed to higher toxicity of wastewater posed on the exogenous bacteria that deal with the included contaminants compared to indigenous bacteria that are inactive towards those contaminants. However, bacteria survived, although of low densities, acquired high resistance against wastewater toxic pollutants [33
Raw wastewater contains huge amount of total coliform (TC) (1.0-9.6x109
CFU/ml). Augmented bacteria showed remarkable ability for TC removal from polluted wastewater. Again, the mixed culture exhibited the highest RE% of TC (99.0%). In contrast control sample showed the opposite trend with continuous increase in the TC starting from 120% after one day to 760% after 7 days indicating the absence of antagonistic bacteria that can kill and assimilate coliform bacteria [35
]. Also, very high fecal coliform (FC) density (1.0-6.9x109
CFU/ml) was recorded in the raw wastewater. However, augmented bacteria showed remarkable ability for FC removal from polluted wastewater with the mixed culture exhibited the highest (almost 100%) removal of FC. Control wastewater FC kept the same trend as for the TC where it recorded continuous increase in the FC ranged between a minimum of 300% and maximum of 800% due to the absence of antagonistic bacteria [37
]. TC that includes both fecal and non-fecal coliform in the treated sample was slightly higher (0.1x109
CFU/ml or 1x108
, i.e. 1000x105
CFU/ml) which means 200 fold higher than the MPL of the TC. However, such increase is mainly composed of non-pathogenic coliform bacteria which represents much lower environmental risk and can be easily removed by the traditional chlorination in a subsequent disinfection stage.
Results of the present study confirmed the ability of the selected bacteria for the removal of the target contaminants including antagonistic effect against pathogenic bacteria (coliform) and thus can be manipulated efficiently to decontaminate polluted systems providing the optimum degradation conditions. Removal of such contaminants was controlled by microbial species, concentration of pollutants in the tested wastewater and finally the contact time between wastewater and the bacterial cells.