Halotolerant Biofilm in Coffee Beans for Phenanthrene Degradation Under Selected Culture Conditions through a Plackett-Burman Experimental Design

Abstract

A halotolerant biofilm was developed on green coffee beans by sequential enrichment in the presence of diesel in order to degrade phenanthrene. Experiments were performed under several combined culture conditions using a Plackett-Burman experimental design. The microorganisms composing the biofilm were identified. These included three fungi, Aspergillus niger, Fusarium solani and Fusarium oxysporum, identified by the Fungal Biodiversity Centre of Holland by macroscopic and microscopic morphological evaluation and by molecular techniques (18s-ITS1-5.8SITS2- 28s rRNA); two yeasts, Candida orthopsilosis and Rhodotorula mucilaginosa; and two bacteria, Pseudomonas putida and Klebsiella variicola, identified by the Mexican Collection of Microbial Cultures (Coleccion Mexicana de Cultivos Microbianos, CINVESTAV) according to colony morphology and biochemical [API 20C AUX and API20 (NE and E), respectively] and molecular (fragments ITS1-5.8S-16s-26s and 16s rRNA, respectively) tests. Each microorganism and biofilm were tested for their tolerance to diesel and salinity conditions and their capacity to degrade phenanthrene; degradation capacity was affected significantly (α<0.05) at low concentrations of NaNO3 and KH2 PO4 and at high concentrations of peptone, CaCO3 and FeSO4 (R2=0.95; C=24.97). Two of the tested conditions, T2 and T5, degraded 91.92% and 89.33%, respectively, of 180 mg/L of phenanthrene. Thus, the biofilms developed from microorganisms of green coffee beans preadapted to salinity conditions presented the metabolic capability to remove phenanthrene from seawater under selective culture conditions.