Batch Culture Enrichment of Indiginous Soil Microorganisms Capable of Catabolizing Creosote Components

Abstract

Batch enrichment cultures for creosote catabolizing microorganisms was carried out using a creosote-contaminated soil as the inoculum. The flasks were separately spiked with phenol, o-cresol, m-cresol, p-cresol, naphthalene, anthracene, phenanthrene, pyrrole, fluorene, pyrene, fluoranthene, chrysene,benzo(a)pyrene and creosote (a complex mixture of about 400 compounds) in concentrations of 50, 100, 500, 1000, 5000, 10 000, 15 000, 20 000, 25 000 and 30 000 mg L-1. The flasks were incubated on a rotary shaker in the dark at 30 °C. Samples for analysis were taken from the flasks every three days for three weeks. Counts of microorganisms were observed to be highest in most cases in 5000 mg L-1 cultures. The pH values were observed to fluctuate between 5 and8 but this did not seem to affect the growth of the organisms except at 50 and 100 mg L-1 in the phenolics and some polycyclic aromatic hydrocarbons (PAHs) were the decreases correlated with decreases in cell counts. The isolates in the first week were mainly of one morphological type in most cultures. In subsequent weeks, the populations became mixed. The higher molecular mass PAHs at higher concentration continued to support only a few types of organisms. Isolates included bacteria, actinomycetes and fungi. The phenolics and naphthalene were more readily removed (up to 100%) from the cultures. Other hydrocarbons removal were between 45 and 83% with the higher molecular mass compounds being most recalcitrant. The decreases in creosote concentration was similar to those in the phenolic compounds and the lower molecular mass compounds up to a concentration of 10 000mg L-1. Decreases in creosote concentration started to decrease from a concentration of 15 000 mg L-1 upward. Subsequent subculturing was observed to enhance the degradative capabilities of the isolates and the further removal of the higher molecular mass compounds. However, this enhanced degradation was much lower in creosote cultures than in the other cultures.