Evaluation of resistance patterns and bioremoval efficiency of hydrocarbons and heavy metals by the mycobiome of petroleum refining wastewater in Jazan with assessment of molecular typing and cytotoxicity of Scedosporium apiospermum JAZ-20

Abstract

Jazan Industrial Economic City (JIEC) is located on the Red Sea coast in the province of Jazan, southwest of Saudi Arabia anchors diverse heavy and secondary industries in the energy, water desalination, petroleum, aluminum, copper, refineries, pharmaceuticals and food manufacturing fields. These various industries generate a large quantity of industrial wastewaters containing various toxicants. The present work represents ecologically beneficial alternatives for the advancement of environmental biotechnology, which could help mitigate the adverse impacts of environmental pollution resulting from petroleum refining effluents. The mycobiome (32 fungal strains) isolated from the industrial wastewater of the refinery sector in Jazan were belonged to five fungal genera including Fusarium, Verticillium, Purpureocillium, Clavispora and Scedosporium with a distribution percentage of 31.25, 21.88, 15.63, 12.50 and 18.75 %, respectively. These isolates showed multimetals tolerance and bioremoval efficiency against a large number of heavy metals (Fe²⁺, Ni²⁺, Cr⁶⁺, Zn²⁺, As³⁺, Cu²⁺, Cd²⁺, Pb²⁺, Ag⁺ and Hg²⁺) along with potent bioremediation activity toward crude oil and the polycyclic aromatic hydrocarbons. Interestingly, the mycobiome resistance patterns obtained against different classes of fungal antibiotics including azole (fluconazole, itraconazole, voriconazole, posaconazole, isavuconazole and ketoconazole), echinocandin (anidulafungin, caspofungin and micafungin) and polyene (amphotericin B) drugs proved the prevalence of antibiotic resistance among the mycobiome of refinery industry in Saudi Arabia is relatively low. The fungal isolate under isolation code JAZ-20 showed the highest bioremoval efficiency against heavy metals (90.8-100.0 %), crude oil (89.50 %), naphthalene (96.7 %), phenanthrene (92.52 %), fluoranthene (100.0 %), anthracene (90.34 %), pyrene (85.60 %) and chrysene (83.4 %). It showed the highest bioremoval capacity ranging from 85.72 % to 100.0 % against numerous pollutants found in a wide array of industrial effluents, including diclofenac, ibuprofen, carbamazepine, acetaminophen, sulfamethoxazole, bisphenol, bleomycin, vincristine, dicofol, methyl parathion, atrazine, diuron, dieldrin, chlorpyrifos, profenofos and phenanthrene. The isolate JAZ-20 was chosen for molecular typing, cytotoxicity assessment, analysis of volatile compounds and optimization investigations. Based on phenotypic, biochemical and phylogenetic analysis, strain JAZ-20 identified as Scedosporium apiospermum JAZ-20. This strain is newly discovered in industrial effluents in Saudi Arabia. Fungal strain JAZ-20 consistently produced various types of saturated and unsaturated fatty acids. the main fatty acids were C_14:0 (1.95 %), iso-C_14:0 (2.98 %), anteiso-C_14:0 (2.13 %), iso-C_15:0 (9.16 %), anteiso-C_15:0 (11.75 %), C_15:0 (7.42 %), C_15:1 (2.37 %), anteiso-C_16:0 (3.4 %), C_16:0 (10.3 %), iso-C_16:0 (9.5 %), C_17:1 (1.36 %), anteiso-C_17:1 (8.64 %), iso-C_18:0 (11.0 %), C_18:0 (3.63 %), anteiso-C_19:0 (3.78 %), anteiso-C_20:0 (2.0 %), iso-C_21:0 (2.44 %), C_23:0 (1.15 %), and C_24:0 (2.17 %). These fatty acids serve as natural and eco-friendly antifungal agents, promoting fungal resistance and inhibiting the production of mycotoxins in the environment. Despite being an environmental isolate, its cytotoxicity was assessed against both normal and cancerous human cell lines. The IC₅₀ values of JAZ-20 extract were 8.92, 10.41, 20.0, 16.5, and 40.0 μg/mL against WI38, MRC5, MCF10A, HEK293 and HDFs normal cells and 43.26, 33.75, and 40.0 μg/mL against liver (HepG2), breast (A549) and cervix (HeLa) cancers, respectively. Based on gas chromatography-mass spectrometry (GC-MS), analysis the extract of S. apiospermum JAZ-20 showed 47 known volatile compounds (VOCs) for varied and significant biological activities. Enhancing the bioremoval efficiency of heavy metals from actual refining wastewater involves optimizing process parameters. The parameters optimized were the contact time, the fungal biomass dosage, pH, temperature and agitation rate.

Keywords: Anti-proliferation; Antibiotic resistant; Bioremoval heavy metals and hydrocarbons; GC/MS analysis; Refinery wastewater; Scedosporium.

Fig. 1

The whole map of Saudi Arabia with Jazan highlighted in the southwestern (a), the map of the Jazan city (b), the study area in Jazan economic city (c) and the refinery industry in the industrial zone of Jazan (d). (a, b, Al-Sheikh et al. [8]; c, d Google maps).

Fig. 2

The bioemoval efficiency (%) of different hazard pollutants from aqueous solutions by the indigenous mycobiome of the refinery wastewater. Each adsorption value is the average value of bioemoval obtained from the isolates of each genus.

Fig. 3

Photomicrograph of colony characteristics (a), microscopic morphology; (b), lactophenol cotton blue magnification × 400) showing solitary and cylindrical conidiophores with graphium morphology of S. apiospermum (b) and scanning electron microscopy (c, the scale is 10 μm) showing cylindrical, thin and septate hyphae, globose to subglobose thick-walled large conidia with a diameter 8–10 × 4–6 μm.

Fig. 4

Phylogenetic tree obtained from the ITS sequence data of the selected isolate JAZ-20 and sequences of reference strains obtained from GenBank, by neighbor-joining method.

Fig. 5

Anti-proliferative activity of S. apiospermum JAZ-20 extract against different human normal cell lines.

Fig. 6

Cytotoxic activity of S. apiospermum JAZ-20 extract against liver, lung and cervix human adenocarcinoma cell lines.

Fig. 7

GC/MS chromatogram of the extract of S. apiospermum JAZ-20 isolated from the industrial wastewater of refinery factories in the industrial area of Jazan.

Fig. 8

Chemical structure of some bioactive volatile compounds detected in the ethyl acetate extract of JAZ-20 fungus.

Fig. 8

Chemical structure of some bioactive volatile compounds detected in the ethyl acetate extract of JAZ-20 fungus.