2006; Steliga et al. 2012), the comparison of existing data is difficult due to the
differences in methodology. The results of Ames test may differ depending on
applied extraction technique, solvent, bacterial strain as well as the presence of
preincubation step (Jadczyk and Kołwzan 2013). The study of Lah et al. (2008)
also suggested that Ames test may not be sensitive to elevated concentrations of trace
elements, which often occur in industrial soils contaminated with TPH, and may
results in misleading conclusions. Standard procedures of Ames test for soil samples
still need to be developed (Jadczyk and Kołwzan 2013).
Microtox
® is another common bioassay for measurement of acute toxicity of soil
eluates (Juteau et al. 2003; Pagnout et al. 2006; Mao et al. 2009; Khudur et al. 2015).
The assay is based on natural or genetically modified bioluminescence ability of
bacterial strains. Aliivibrio fischeri, due to natural ability of bioluminescence is a
standard organism for Microtox
® bioassay. However, as it is a marine bacterium
doubts are arising due to application as a reference to contaminated soil. In the study
of Bundy et al. (2004), the response of A. fischeri was compared to common soil
bacterium, Pseudomonas putida, modified with a plasmid encoding bioluminescence. Obtained results showed opposite reaction of both strains after contact with
leachates of soil spiked with paraffin and motor oil. Differences in the response of
both strains for the same contaminant highlight the importance of bioindicator
selection for standard methods. However, considering the importance of existing
literature, Microtox
® is still a useful tool for soil toxicity analysis.
Both abovementioned bioassays are common procedures for soil ecotoxicity
monitoring. However, the leachates do not reflect exactly the composition of the
soil matrix, which may also influence toxicity. Mechanism of TPH toxicity is based
on partitioning into cell membranes, thus, the effect on the cell toxicity may not be as
visible under bacterial toxicity assay due to limited contact with the contaminant. In
conclusion, a bioassay based on single species is not expected to give reliable results
as the response of each species depends on specific sensibility to contaminant
(Bundy et al. 2004). Thus, to enhance the relevance of ecotoxicological evaluation,
bioassays employing plants or invertebrates may be used together with bacterial
response assays.
5.4.3.2 Phytoassays
Phytoassays are performed by direct planting of test species on contaminated soil or
by tests on soil leachates. For TPH-contaminated soil contact tests are more effective
tool for ecotoxicity evaluation in comparison with tests performed on soil leachates
(Hubálek et al. 2007). The test performed on soil eluates are faster and easier to
perform but also less sensitive and may result in inappropriate estimation of soil
toxicity (Hubálek et al. 2007). Direct germination tests may provide information
about germination rate, plant fresh, and dry biomass as well as the length of root and
shoot. The results may be expressed as Germination Index or Growth Inhibition
Index (Graj et al. 2013).
5 Potential Use of Waste-to-Bioenergy By-Products in Bioremediation of Total. . .
271
differences in methodology. The results of Ames test may differ depending on
applied extraction technique, solvent, bacterial strain as well as the presence of
preincubation step (Jadczyk and Kołwzan 2013). The study of Lah et al. (2008)
also suggested that Ames test may not be sensitive to elevated concentrations of trace
elements, which often occur in industrial soils contaminated with TPH, and may
results in misleading conclusions. Standard procedures of Ames test for soil samples
still need to be developed (Jadczyk and Kołwzan 2013).
Microtox
® is another common bioassay for measurement of acute toxicity of soil
eluates (Juteau et al. 2003; Pagnout et al. 2006; Mao et al. 2009; Khudur et al. 2015).
The assay is based on natural or genetically modified bioluminescence ability of
bacterial strains. Aliivibrio fischeri, due to natural ability of bioluminescence is a
standard organism for Microtox
® bioassay. However, as it is a marine bacterium
doubts are arising due to application as a reference to contaminated soil. In the study
of Bundy et al. (2004), the response of A. fischeri was compared to common soil
bacterium, Pseudomonas putida, modified with a plasmid encoding bioluminescence. Obtained results showed opposite reaction of both strains after contact with
leachates of soil spiked with paraffin and motor oil. Differences in the response of
both strains for the same contaminant highlight the importance of bioindicator
selection for standard methods. However, considering the importance of existing
literature, Microtox
® is still a useful tool for soil toxicity analysis.
Both abovementioned bioassays are common procedures for soil ecotoxicity
monitoring. However, the leachates do not reflect exactly the composition of the
soil matrix, which may also influence toxicity. Mechanism of TPH toxicity is based
on partitioning into cell membranes, thus, the effect on the cell toxicity may not be as
visible under bacterial toxicity assay due to limited contact with the contaminant. In
conclusion, a bioassay based on single species is not expected to give reliable results
as the response of each species depends on specific sensibility to contaminant
(Bundy et al. 2004). Thus, to enhance the relevance of ecotoxicological evaluation,
bioassays employing plants or invertebrates may be used together with bacterial
response assays.
5.4.3.2 Phytoassays
Phytoassays are performed by direct planting of test species on contaminated soil or
by tests on soil leachates. For TPH-contaminated soil contact tests are more effective
tool for ecotoxicity evaluation in comparison with tests performed on soil leachates
(Hubálek et al. 2007). The test performed on soil eluates are faster and easier to
perform but also less sensitive and may result in inappropriate estimation of soil
toxicity (Hubálek et al. 2007). Direct germination tests may provide information
about germination rate, plant fresh, and dry biomass as well as the length of root and
shoot. The results may be expressed as Germination Index or Growth Inhibition
Index (Graj et al. 2013).
5 Potential Use of Waste-to-Bioenergy By-Products in Bioremediation of Total. . .
271