Methylotrophs and Hydrocarbon-Degrading Bacteria Are Key Players in the Microbial Community of an Abandoned Century-Old Oil Exploration Well

In this work, we studied the microbial community and the physicochemical conditions prevailing in an exploratory oil well, abandoned a century ago, located in the Cahuita National Park (Costa Rica). According to our analysis, Cahuita well is characterized by a continuous efflux of methane and the presence of a mixture of hydrocarbons including phenanthrene/anthracene, fluoranthene, pyrene, dibenzothiophene, tricyclic terpanes, pyrene, sesquiterpenes, sterane, and n-alkanes. Based on the analysis of 16S rRNA gene amplicons, we detected a significant abundance of methylotrophic bacteria such as Methylobacillus (6.3-26.0% of total reads) and Methylococcus (4.1-30.6%) and the presence of common genera associated with hydrocarbon degradation, such as Comamonas (0.8-4.6%), Hydrogenophaga (1.5-3.3%) Rhodobacter (1.0-4.9%), and Flavobacterium (1.1-6.5%). The importance of C1 metabolism in this niche was confirmed by amplifying the methane monooxygenase (MMO)-encoding gene (pmo) from environmental DNA and the isolation of two strains closely related to Methylorubrum rhodesianum and Paracoccus communis with the ability to growth using methanol and formate as sole carbon source respectively. In addition, we were able to isolated 20 bacterial strains from the genera Pseudomonas, Acinetobacter, and Microbacterium which showed the capability to grow using the hydrocarbons detected in the oil well as sole carbon source. This work describes the physicochemical properties and microbiota of an environment exposed to hydrocarbons for 100 years, and it not only represents a contribution to the understanding of microbial communities in environments with permanently high concentrations of these compounds but also has biotechnological implications for bioremediation of petroleum-polluted sites.