Two camps exist with regard to the chief rate-limiting factor in biodegradation: 1) mass transfer versus 2) microbial activity. This work will not address mass transfer quantitatively. A qualitative evaluation is given in section 2.6 (“factors affecting contaminant
bioavailability”).
The microbial factors affecting degradation rates include, but are not limited to, prior exposure, the presence of a second organic substrate, the accumulation of dead-end or toxic metabolites, the presence and numbers of specific bacterial or fungal strains, the presence of inducers, and the rhizosphere effect.
2.7.1 Prior exposure Prior exposure of microbial populations to a contaminant leads to
enhanced initial rates of degradation most likely because of a diminished lag period during the initial phase post-exposure. While the enzymes responsible for PAH degradation may or may not be constitutive, repeated exposure to a compound leads to an acquired adaptive capability. Johnsen and Karlsen (2005) found that soils near industrialized areas contained enough pyrene, even at low levels, to sustain degradative genes within the indigenous
community. However, prior exposure does not always lead to increased rates of degradation, depending on how the studies are performed. Smith et al. (1999) added known PAH
degraders to crude-oil contaminated soils and found preferential degradation of other oil substituents besides PAH. Viñas et al. (2002) also found that an enrichment of degraders from specific fractions of oil (e.g. aromatic) did not make them more efficient degraders of that particular fraction when augmented into an oil-medium emulsion.
2.7.2 Degrader numbers The measure of PAH-degrader numbers as a reflection of PAH
number of degraders leads to increased degradation rates (Gentry et al., 2003; Del Panno et al., 2005), others have found a poor correlation between numbers and degradation
(Carmichael and Pfaender, 1997). Huesemann et al. (2002) showed a decrease in the ratio of hydrocarbon degraders to total heterotrophs over time for most of the soils they tested. Furthermore, this increase in numbers may not always be attainable or sustainable. For example, energy resulting from the initial phase of rapid degradation of a PAH mixture may be needed for cell maintenance rather than growth, once the usable carbon (e.g. low-
molecular-weight PAH) has been depleted or is no longer available. It may be that functional redundancy or cometabolic effects rather than numbers play a more significant role.
2.7.3 Substrate competition Competition for the enzyme active site by two PAHs may
account for reduced rates of degradation of one of the compounds (Bouchez et al., 1995, 1996) and was cited as the reason for a decrease in phenanthrene degradation by a mixed culture when a second PAH was present (Stringfellow, 1994) and a decrease in the pyrene degradation rate by M. flavescens in the presence of fluoranthene (Dean-Ross et al., 2002). Bouchez et al. (1999) found that the rate of mineralization decreased as the oxygen/carbon ratio decreased, for example with the addition of a second carbon source. Pyrene was cometabolized in the presence of anthracene as the primary carbon source but not when fluoranthene or phenanthrene was present.
2.7.4 Presence of inducers Salicylate is the most commonly studied inducer of PAH
metabolism. It is an intermediate in many degradation pathways, and its production induces the lower pathway for naphthalene degradation. Its effects do not appear to be consistent and may depend on the PAH, the salicylate concentration, and the organisms present. The
benz[a]anthracene, chrysene, and benzo[a]pyrene by Pseudomonas saccharophila P15 in liquid culture (Chen and Aitken, 1999). However, Carmichael and Pfaender (1997) were unable to show any effect of its addition on phenanthrene and pyrene degradation in soil microcosms. Repeated additions of salicylate induced the greatest extent of pyrene mineralization in soil microcosms as compared to a single spike (Vanderford, 2001).
2.7.5 Presence of metabolites “Dead-end” metabolites, or those that are degraded no
further, have been shown to accumulate in some systems during the initial period of fast cell growth and compound degradation. Some of these have been deemed toxic to down-stream metabolizers. Kazunga and Aitken (2000) showed that the formation of cis-4,5-dihydro-4,5- dihydroxypyrene (PYRdHD) by bacterial isolates from a PAH-contaminated soil caused inhibition of phenanthrene degradation by some of the strains, while both PYRdHD and pyrene-4,5-dione (PYRQ) resulted in the inhibition of benzo[a]pyrene degradation by others. This latter finding was supported by Vanderford (2001) who used Microtox assays to show that PYRQ caused a decrease in the ATP-dependent production of luciferase by test
organisms at levels well below its aqueous solubility. Although the above metabolites result from oxidation at bay- and K-regions of the PAH, others have shown that attack at non-bay- and non-K-regions may also result in toxic, dead-end metabolites (Kim et al., 2005).
Increased degradation of a PAH mixture was reported for a consortium of PAH degraders as compared to a mixed, but defined, co-culture of bacteria, possibly due to a build-up of toxic metabolites that the defined culture was ill-suited to degrade (Bouchez et al., 1999).
2.7.6 Presence of fungi The presence of fungi in a mixed culture has been shown to have
differing effects. As mentioned earlier, fungi may play a role in the initial oxidation step of PAHs, in their incorporation into humic matter, or in the formation of toxic metabolites.
PAHs may adsorb to fungal mycelia, leading to a decrease in availability to bacterial degraders. Hyphae also play an important role in the formation, maintenance, and
destruction of soil aggregates. This can lead to the spatial isolation of some bacteria, thereby decreasing their accessibility to contaminants. The presence of organic-rich matter in soils was shown to stimulate ligninolytic fungi and, as a result, enhanced rates of PAH
degradation. Fungi in organic-poor soils had an inhibitory effect on degradation, possibly due to the lower production of oxidative enzymes by mycelia under these conditions and the repression of bacterial degraders. In either event, the presence of fungi may alter the
evolving bacterial community (Gramss et al., 1999).
2.7.7 Plant presence Some have studied the effects of plant residues (“the rhizosphere
effect”) on bacterial populations in contaminated soils. Joner et al. (2002) measured
enhanced degradation of 3- and 4-ring PAHs after the addition of phosphorus, nitrogen, and root exudates and the increase in the ratio of PAH-degrading bacteria to total heterotrophs. Corgié et al. (2004) found that different bacterial communities were selected as a function of the distance to plant roots and that those located closest to roots had greater phenanthrene degrading abilities, possibly due to stimulation by root exudates.