Evaluation of crude oil biodegradation using mixed fungal cultures

The use of potent fungal mixed cultures is a promising technique for the biodegradation of crude oil. Four isolates of fungi, namely, Alternaria alternata (AA-1), Aspergillus flavus (AF-3), Aspergillus terreus (AT-7), and Trichoderma harzianum (TH-5), were isolated from date palm soil in Saudi Arabia. The mixed fungal of the four isolates have a powerful tool for biodegradation up to 73.6% of crude oil (1%, w/v) in 14 days. The fungal consortium no. 15 containing the four isolates (1:1:1:1) performed significantly better as a biodegradation agent than other consortium in a variety of environmental factors containing crude oil concentration, incubation temperature, initial pH, biodegradation time and the salinity of the medium. The fungal consortium showed better performance in the biodegradation of normal alkanes (n-alkanes) than that of the polycyclic aromatic hydrocarbons (PAHs); the biodegradation efficiency of normal alkanes of the fungal consortium (67.1%) was clearly high than that of the PAHs (56.8%).


Unfunded studies
Enter: The author(s) received no specific funding for this work.   Yes -all data are fully available without restriction

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Many researchers are improving fungal consortia for the complete degradation of such 40 hydrocarbons. In these consortia, three strategies of biodegradation are represented: (1) The target 41 hydrocarbon compound is used as a carbon source by fungi; (2) the target hydrocarbon compound is 42 enzymatically attacked but is not used as a carbon source (cometabolism); (3) the target compound is 43 not metabolized at all but is absorbed and accumulated within the fungi (bioaccumulation). Although 44 fungi participate in all three strategies, they are more effective at cometabolism and bioaccumulation 45 than at using hydrocarbons as sole carbon sources [9]. Fungi can provide powerful and unique of     The four tested fungal isolates were cultured individually in potato dextrose broth medium 77 (PDB) (20 g of potato and 2 g of dextrose) in 250 mL conical flasks, to which 0. At the same time, the four fungal isolates (both individually and in combinations), which were 98 co-cultivated with crude oil as mentioned above, were also tested for enzyme activity. The enzyme 99 level in the medium was assayed for control and treatment after 2 weeks of incubation. At 10,000 rpm 100 and 4 ° C, the cultures of fungi were centrifuged and the enzyme level was determined. Cellulase    The pH of the MSM medium was set to 6, 7, 8, and 9 with 1 mol / L of NaOH and HCl and 119 solution, respectively. The medium included the crude oil and, the fungal consortium (5 mL) was 120 inoculated and the biodegradation efficiency was estimated at 20 ° C after 7 days of fungal culture.

Analysis of crude oil degradation by GC-MS
The best fungal consortium has been tested for its ability to degrade crude oil by studying its 131 structure before and after the fungal activity. This experiment was applied, as mentioned in section 132 2.4. At the time of the GC-MS analysis, the sample of crude oil degradation mixed with diethyl ether 133 at a ratio of 1:1. The samples were agitated for 1 min, and the organic layer was released to separate 134 after 5 min. At this moment, collecting 1 mL of the supernatant (crude oil degradation) to a 1.5 mL 135 Eppendorf tube, and centrifuged at 3000 rpm for 3 min. 1 ml of the sample was put in a glass vial.

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The crude oil extracted from sample was evaluated and used a flame ionization detector via a Perkin-   Table 2. Based on the cluster analysis of BLAST and the ITS sequence for each fungal 149 isolate proved that isolate AA-1 was 99% similar to A. alternata (LC164851). Using the same method,

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The screening technique was dependent on the changing of color of the fungal isolates treated 159 with the redox indicator (Table 3) although all of the fungal isolates were confirmed to possess the 160 ability to degrade crude oil to differing levels, the best isolates were AF-3 and TH-5, followed by AT-     The accumulation of contaminant hydrocarbons influences fungal biodegradation in the 238 bioremediation process, and too high an accumulation contributes to a very decline in the 239 performance of fungal biodegradation. As observed in Table 9, the crude oil concentration  for improved growth of the majority of bacteria is between pH 6.5 and 8.5, whereas the pH condition 292 required for yeast and fungi is between pH 4.5 and 5.5. Thus, an optimal pH range for biodegradation 293 of crude oil might be considered between pH 6.0 and 8.5. The pH can be highly variable and must be 294 taken into consideration when improving biodegradation treatment methods. The environmental pH 295 influences physiological processes such as transport through cell membrane and catalytic reaction 296 balance well as enzyme activities [44]. The pH range for improved growth of most bacteria is between 297 pH 6.5 and 8.5, in the pH 4.5-5.5 and in the pH 4.5-6.5 range for fungi. Therefore, an ideal pH range 298 may be considered between pH 6.0 and 8.5 for the microbial biodegradation of crude oil [45]. MnP 299 activity for Aspergillus niger consistently decreased in the range of pH 5.5-8.5, and was optimum at 300 pH 5.5. MnP activity for Penicillium freii was lowest at pH 7.0 and optimum at pH 8.5 [46]. MnP 301 activity for Aspergillus terreus was optimum at an alkaline pH of 12.5 and maintained its stability 302 between the pH 11.0 and 12.5 [47]. The variation in the optimum pH of the enzyme activity between 303 fungal isolates used in many of the cited studies could be due to the difference in substrates used 304 or/and the strength of the isolate.

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The experiments of Crude oil biodegradation were conducted at salinity ranging from 2 % to 8 307 % to confirm the effectiveness of a mixture of three or four fungal isolates together as a consortium 308 based on a synergistic effect that can tolerate wider salinity changes, and the findings are shown in