Microbiological contamination in petroleum-based fuels has been exacerbated with the addition of fatty acid methyl esters to diesel fuels. Consequences of microbiological contamination of these fuels can lead to degraded fuels, fouling and clogging of infrastructure, and potentially lead to microbiologically influenced corrosion (MIC) from the formation of organic acids as these microbes metabolize the fuel components. Additionally, operators are typically unaware of any potential contamination in their fuel tanks as the formation of biofilms can interfere with current technology designed to alert them of ongoing problems. As part of this dissertation, the fungal isolates Paecilomyces AF001 and Wickerhamomyces SE3 were characterized as being capable of degrading B20 biodiesel and using it as the sole carbon and energy source. The metabolism of B20 biodiesel led to an acidification of the medium and caused an increase in pitting corrosion and generalized corrosion on carbon steel. Additionally, this research has provided evidence that corrosion risks in contaminated fuel storage tanks are greatest at the interface of the fuel and any water that becomes entrapped in the fuels. Prior to this research, limited information was known about the microbiological communities in ultra-low sulfur diesel (ULSD) and B20 biodiesel. This research expands the knowledge of microbial communities in fouled fuels by analyzing contaminated fuels from 106 fuel tanks at 17 military bases across the continental U.S. This research has demonstrated the bacterial communities in contaminated fuels are far more diverse than fungal communities in fuels when they are present. When fungal contamination occurred in fuels it was primarily composed of the filamentous fungal family Trichocomaceae. Fuel composition of B5 ULSD and B20 biodiesel was determined and used to correlate microbial community families to the fuel components. The same problematic fungal isolates Paecilomyces AF001 and Wickerhamomyces SE3 used to evaluate corrosion of carbon steel when grown on B20 were used to determine if the correlations predicted by RDA analysis were accurate. Trichocomaceae (representative isolate Paecilomyces AF001) had positive correlations with fuels containing more palmitoleic acid methyl ester and the fungal family Debaryomycetaceae (representative isolate Wickerhamomyces SE3) had a positive correlation with increases in pentadecanoic acid methyl esters in fuels. Both isolates were grown on these substrates to determine their ability to utilize them as a sole carbon and energy source. Paecilomyces AF001 was able to grow on palmitoleic acid methyl ester and was unable to grow on pentadecanoic acid methyl esters while Wickerhamomyces SE3 was able to grow on both substrates. Fungal families are less diverse than bacterial families in contaminated fuels and were primarily present when contamination occurred. Due to this, the fungal family Trichocomaceae which was present at many contaminated fuel storage tanks, was selected for enzymatic and transcriptomic analyses on B5 ULSD and B20 biodiesel as the sole carbon and energy sources. Paecilomyces AF001, a member of the Trichocomaceae family, has already been shown to be able to utilize hydrocarbons and FAME (Fatty Acid Methyl Ester) components in fuels and leads to increased corrosion risks. Transcriptomics was done to see any differences in metabolic utilization of genes associated with the metabolism of hydrocarbons and FAME. Paecilomyces was able to grow on both B5 ULSD and B20 biodiesel. Transcripts associated with hydrocarbon degradation, such as mono and dioxygenases, were higher than those seen when this fungus grew in B20 biodiesel. Additionally, lipase activity and transcripts associated with lipase genes were observed in both fuel types; however, more lipase activity and transcripts were found when Paecilomyces AF001 was grown on B20 fuel instead of B5 ULSD. Understanding how Paecilomyces AF001 grows on different fuel types can lead to the development of biosensors that can help operators detect contamination in their tanks sooner and hopefully lead to less costs associated with remediating contaminated tanks. Overall, this work has demonstrated that microbial contamination of B5 ULSD and B20 is a rampant problem across the U.S. This work has linked the filamentous fungus Paecilomyces AF001 to increased corrosion risks to carbon steel when grown on B20 biodiesel, demonstrated that the fungal family Trichocomaceae (representative isolate Paecilomyces AF001) is a predominant fouler when storage tanks are contaminated, and demonstrates that this organism transcribes different genes associated with fuel metabolism based on if this organism is grown on B5 ULSD or B20 biodiesel.