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Article

Porcellio scaber Latr. and Lumbricus terrestris L.—PAHs Content and Remediation of Long-Term Aging Soil Contamination with Petroleum Products during a Single- and Two-Species Experiment

by
Janina Gospodarek
1,*,
Przemysław Petryszak
2,
Alina Kafel
3 and
Iwona B. Paśmionka
1
1
Department of Microbiology and Biomonitoring, University of Agriculture, al. A. Mickiewicza 21, 31-120 Krakow, Poland
2
Unit of Biochemistry, Institute of Plant Biology and Biotechnology, Faculty of Biotechnology and Horticulture, University of Agriculture in Krakow, al. 29-Listopada 54, 31-425 Krakow, Poland
3
Department of Animal Physiology and Ecotoxicology, University of Silesia, Bankowa 9, 40-007 Katowice, Poland
*
Author to whom correspondence should be addressed.
Energies 2022, 15(21), 7835; https://doi.org/10.3390/en15217835
Submission received: 20 September 2022 / Revised: 19 October 2022 / Accepted: 20 October 2022 / Published: 22 October 2022
(This article belongs to the Section B: Energy and Environment)
Browse Figures
Versions Notes

Abstract

:
The presence of earthworms in soil contaminated with petroleum products (PPs) increase the rate of decomposition. The use of more than one animal species simultaneously in soil remediation could accelerate the process. However, the effects may be different when long-term aging soil contamination exists during which toxic by-products can be formed. This study evaluated the effect of soil contamination with PPs (petrol, diesel fuel, and used engine oil) carried out 12 and 24 months earlier on the life parameters of Lumbricus terrestris L. during single- and dual-species breeding with Porcellio scaber Latr. The changes in the content of total petroleum hydrocarbons (TPH) in the soil were also measured, as were the levels of polycyclic aromatic hydrocarbon (PAH) accumulation in the bodies of the test animals. Survival of earthworms cultivated separately (single-species experiment) in soil contaminated with diesel fuel 12 months earlier significantly decreased, while body mass gain was not significantly changed under the influence of tested PPs. Breeding of L. terrestris together with P. scaber contributed to significant mortality of earthworms regardless of treatments, while woodlice showed much greater resistance to PPs. Occurrence of both animals in soil contaminated with engine oil 24 months earlier resulted in a significant reduction in TPH after 4 weeks (by 29% compared to the initial soil). The content of PAHs in the tissues of L. terrestris was higher than in P. scaber, and the highest amounts of PAHs were found when earthworms were exposed to engine oil. Use of L. terrestris together with P. scaber can be considered a promising scenario for increasing the effectiveness of remediation of soils contaminated with petroleum products, however, further research is needed to establish conditions preventing excessive mortality of earthworms in such a layout.

1. Introduction

Petroleum products (PPs), being substances commonly used around the world, are a source of soil contamination with a number of biohazardous compounds. Contamination most often occurs at PP processing and storage sites as well as during transportation [1]. Soil organisms are a group highly exposed to such pollution, often responding with changes in growth, fertility, increased mortality, or physiological changes [2]. Some of them at the same time may be involved in the natural breakdown of pollutants [3,4,5].
Porcellio scaber Latr. is a common representative of invertebrates, playing an important role in the decomposition of organic matter mainly by pre-crushing and mixing it with soil particles but also by secreting specific chemical substances, such as enzymes (e.g., cellulases) or surfactants, which are produced mainly by endosymbiotic bacteria belonging to the Bacillus, Pseudomonas, or Staphylococcus genera inhabiting the digestive tract or hepatopancreas [6,7,8,9,10]. There is a close relationship between the presence of microorganisms responsible for organic compound decomposition processes and the feeding preferences of P. scaber. In particular, P. scaber showed a positive olfactory reaction against microbial metabolites from cellulose digestion [11]. On the other hand, the presence of P. scaber has a positive effect on microbial biomass and respiration [8]. Concurrently, P. scaber was shown to be quite resistant to short (4 weeks) contact with PPs-contaminated soil, manifested by high survival rate and undisturbed weight gain [12].
The presence of earthworms in soil contaminated with PPs can accelerate the rate of decomposition [3,13,14]. This is achieved through increased soil aeration by earthworm activity, increased (also due to this) microbial activity, and increased availability of hydrocarbons to microorganisms through bioturbation. Lumbricus terrestris L. was found to be the most effective in enhancing oil degradation among the three earthworm species tested (Eisenia fetida Savigny, Allolobophora chlorotica Savigny, and Lumbricus terrestris L.) [3]. In addition, L. terrestris has been used to assess the remediation needs of land contaminated with petroleum substances [15], and although it is not a species included in standard ecotoxicological tests [16], many authors emphasize its usefulness in biomonitoring [17,18,19].
Polycyclic aromatic hydrocarbons (PAHs) are some of the most dangerous components of petroleum substances due to the fact that they can be accumulated in the bodies of organisms exposed to contact with PPs and can also enter downstream in the food chain. P. scaber, taking food contaminated with certain PAHs, may metabolize some of them (e.g., pyrene) while accumulating others [20,21,22,23,24]. However, when exposed only through contact with PP-contaminated soil (and not through food), PAH accumulation has been shown to be quite low [12]. In contrast, earthworms exposed to soil contamination with PAHs accumulate these compounds in varying amounts depending on the species as well as on the type of PAH [25,26].
Most of the studies to date evaluating the effectiveness of invertebrates in accelerating the process of natural remediation of PP-contaminated soil were carried out on the soil contaminated immediately prior to the experiment. However, little is known about the effect of long-term aging soil contamination on the aforementioned abilities of invertebrates as well as on the condition of these animals themselves. The process of natural remediation of PP-contaminated soil takes place with the participation of soil microorganisms that degrade toxic compounds. However, during this process, toxic microbial by-products, such as oxidized derivatives of aliphatic and mono- and polyaromatic hydrocarbons (e.g., cis-4,5-dihydro-4,5-dihydroxypyrene or pyrene-4,5-dione) can also be formed, which may increase cytotoxicity and blockage of enzyme induction [27,28,29].
The use of more than one animal species simultaneously in soil remediation has also been undertaken quite rarely in previous studies. Evaluation of the effectiveness of remediation of soil contaminated with crude oil using two-species breeding of earthworms (L. terrestris and Eudrillus euginae, Kinberg) showed higher TPH loss when both earthworm species were bred separately [30]. On the other hand, Almutairi [14], using two species of earthworms, i.e., Eisenia fetida and L. terrestris, showed higher TPH removal efficiency in Kuwaiti oil-contaminated sand when both species occurred simultaneously than when they were used separately. However, the available literature lacks data on the effects of mixed breeding using animals from more distant systematic units and with different lifestyles, diets, etc. (as represented by earthworms and woodlice).
Research on the use of invertebrates in the remediation of petroleum contaminants has mostly focused on contaminants with crude oil [3,13,30,31,32]. Only a few deal with petroleum products. Specifically, Fernandez et al. [33] studied the degradation of diesel fuel in soil in artificially assembled microcosms during a 180-day experiment, showing higher degradation efficiencies using earthworms compared to phytoremediation using Festuca arundinacea and Trifolium pretense. While crude oils can vary greatly in terms of their chemical composition (different amounts of carbon number C8–C40, different concentrations of asphaltenes, resins, wax, mechanical impurities) [34], petroleum products have a strictly defined composition and properties that are standardized during the production process, and that information is publicly available in the material safety data sheets. Petrol is a complex mixture of volatile hydrocarbons containing paraffins, naphthenes, olefins, and aromatic hydrocarbons which contain between C4 and C12 atoms in the molecule. Diesel fuel is a mixture of hydrocarbons middle distillates, made between C10 and C28 atoms and may also contain fatty acid methyl ester (FAME). Engine oil is a mixture of mineral and synthetic base oils and enriching additives. It is also worth noting that spills of petroleum products (especially diesel fuel) are almost as common as spills of crude oils [35].
Given the information mentioned above, the novel aspects of our investigation would be: firstly, using animals from distant systematic units (earthworms and woodlice) together to test the possibility of increasing the effectiveness of total petroleum hydrocarbon (TPH) removal (as far as we know, this has not been studied yet); secondly, evaluating the impact of long-term aging soil contamination (most studies look at soil contaminated immediately prior to the experiment), and finally, assessing the influence of petroleum products—most of the research on this topic deals with crude oils. Therefore, the objectives of the study were: (a) to evaluate the effect of soil contamination with PPs (petrol, diesel fuel, and used engine oil) carried out 12 and 24 months earlier on the life parameters (survival, body mass change) of L. terrestris during single- and dual-species breeding with P. scaber; (b) evaluation of changes in the content of TPH in the soil during one- and two-species breeding; (c) analysis of the level of polycyclic aromatic hydrocarbon (PAH) accumulation in the bodies of the test animals.

2. Materials and Methods

2.1. Experimental Setup

The soil used in the research (loamy sand, pH in H2O = 7.12, Ctotal = 1.04%) was obtained from a long-term experiment conducted under field conditions (Poland; 50.0815° N, 19.84730° E). The experiment concerned the effects of soil contamination with PPs (petrol, used engine oil, and diesel fuel) in the amount of 6 g of each PP per 1 kg of dry soil mass (i.e., typical concentrations of petroleum in moderately contaminated soils) on soil and epigeic fauna, as well as plants grown in contaminated soil. A detailed description of this experiment is included in a previous paper [36].
Single- (L. terrestris) and two-species (L. terrestris + P. scaber) breeding of animals was carried out for a period of 4 weeks under constant conditions at 22 °C. Individuals of P. scaber were caught near the location of the field experiment but from an uncontaminated area. L. terrestris was purchased from a commercial sport fishing supply. All test animals were acclimated to the laboratory conditions for several weeks before the experimental breeding was carried out.
The following experimental groups were formed:
Control—breeding was conducted on soil without the addition of contaminants;
P—breeding was conducted on soil contaminated with lead-free petrol;
EO—breeding was conducted on soil contaminated with used engine oil;
DF—breeding was conducted on soil contaminated with diesel fuel;
Each week of the experiment, the number of live individuals was recorded, and in the case of single-species breeding, their body weight was also recorded.
Single-species breeding of L. terrestris was conducted on soil taken from the experiment 12 months after soil contamination with PPs, while two-species breeding of L. terrestris + P. scaber was conducted on soil taken after 24 months. The breeding containers each contained 4 L of soil air-dried and sieved through a 1.5 mm sieve and mixed with water. The soil prepared in this way was mixed weekly with air-dried and powdered horse manure (4 g/container). In turn, the woodlice were supplied daily with moistened previously dried leaves of Acer sp. collected in the previous season from a pollution-free area. The soil was sprinkled with water every other day to maintain adequate moisture. In each container, 18 specimens of earthworms with well-developed clitellum were maintained in the case of a single-species breeding of L. terrestris. In the case of two-species breeding, 15 individuals of L. terrestris and 30 individuals of P. scaber were kept in each container. The breeding was carried out in 3 replicates.
During and after breeding (i.e., after two and four weeks of the experiment in the case of L. terrestris and after four weeks in the case of P. scaber), animals were sampled for analysis for PAH content in their bodies. In turn, the experimental soil was analyzed for TPH content. In addition, in the case of two-species breeding, the TPH content of soil kept under the same conditions but without the presence of animals was analyzed.

2.2. Analysis of the Concentration of Petroleum-Derived Compounds in the Soil

2.2.1. Determination of Dry Soil Matter

Porcelain crucibles, in which the dry mass of the soil was determined, were dried at 105 °C for approx. 12 h before the planned measurement. Porcelain dishes were weighed with an accuracy of 0.001 g, and then about 10 g of soil samples were transferred to them. Then the crucibles were incubated at 105 °C for 5 h. After this time, the crucibles were cooled in a desiccator for 1 h and then weighed again. After that, the percentage of dry matter was determined. The presented results of the soil dry matter content are the average of two independent determinations.

2.2.2. Determining Total Petroleum Hydrocarbons Content (TPH) in Soil Samples

Approximately 10–11 g of soil was weighed and placed into closed plastic containers. After the addition of 1 cm3 of 18% hydrochloric acid solution (obtained by mixing concentrated hydrochloric acid with distilled water in the ratio of 1:1; v/v), samples were intensively mixed with glass rods. In order to improve the extraction of TPH, the soil samples were dried, about approximately 12.5 g of anhydrous magnesium sulfate was added to each of them, and they were intensively mixed again. Anhydrous magnesium sulfate was obtained by drying the salt at 105 °C for at least three days before the planned determination. The samples prepared this way were tightly closed and left for approximately 12 h for drying. Before extraction, the soil samples were quantitatively transferred to extraction thimbles (Cellulose Extraction Thimbles, GRADE 603; Sigma-Aldrich, Cat. #: Z612456).
The determination of TPH was performed using the gravimetric method according to the Polish standard PN-C-04573-01:1986 [37] with some further modifications. Erlenmeyer flasks were dried for 12 h in an incubator (temp. 105 °C). Then, they were cooled in a desiccator for 1 h. After this time, the flasks were weighed on an analytical balance with an accuracy of 0.00001 g. The extraction thimbles containing the previously prepared soil samples were transferred to the Soxhlet apparatus (150 cm3) and combined with Erlenmeyer flasks and reflux condensers. The extraction, consisting of repeated washing cycles of the soil samples with an organic solvent, was carried out using petroleum ether (Petroleum Benzine Boiling Range 40–60; Sigma-Aldrich, Cat. #: 32299, St. Louis, MI, USA). Each extraction lasted 6 h, which corresponded to approx. 40 cycles of washing soil samples with an organic solvent. After the extraction was completed, the TPH-containing extracts were concentrated to a post-extraction residue at the bottom of the flasks. The flasks were then dried at 105 °C for 1.5 h to remove residual petroleum ether. After this time, the flasks were transferred to a desiccator, cooled for 1 h, and then reweighed on an analytical balance. The weight of TPH was determined as the difference between the final and initial mass of the Erlenmeyer flasks. The concentration of total petroleum hydrocarbons was calculated based on the obtained results and the previously determined soil dry matter. The determinations were performed in 2 independent repetitions, and the final result obtained is their mean value.

2.3. Analysis of PAHs Content in Animal Tissues

Hexane extracts from animal bodies containing PAH were prepared by weighing 1.0 to 4.0 g of animal tissue into a glass extraction thimble. Then, 1 mL of 1-methylchrysene solution was added and used as the internal standard, followed by 10 mL of hexane. Animal material was homogenized using a high-speed laboratory homogenizer (approx. 10,000 rpm). Extraction consisted of 3 homogenization cycles of 1 min each. After the homogenization cycle was completed, the hexane extract was decanted, and the remaining animal material was again refilled with 15 mL of solvent. In order to obtain clear supernatants, after completion of homogenization, the extracts were centrifuged (3000 rpm; 4 °C, 10 min.). The centrifuged extracts were then transferred quantitatively to ground-glass conical flasks. Finally, the centrifuge tubes were washed with hexane and added to the previously pooled extracts. The pooled extracts were concentrated to dryness in a vacuum evaporator, and then the residue was suspended in 1 mL of hexane and stored for further analysis (−80 °C).
The qualitative and quantitative analysis of the polycyclic aromatic hydrocarbons in the hexane extracts obtained from the animal samples was performed based on gas chromatography and mass spectrometry (GC-MS). For this purpose, a Shimadzu GC—17A ver. 3 equipped with a Shimadzu QP-5000 mass spectrometer was used. The separations were carried out using SLB-5ms capillary column (Supelco, Bellefonte, PA, USA; parameters—60 m × 0.25 mm × 0.25 µm). The capillary column temperature program was as follows: 50 °C (held for 2 min), then the temperature was increased to 330 °C (5 °C/min) and held for 12 min. The injector and linker temperatures were set to 335 °C and 330 °C, respectively. The total separation time was 70 min. The carrier gas’s linear velocity (helium 5.0) was 25 cm/min. Samples (1 µL) were injected using a precision microsyringe (Hamilton) and the Shimadzu AOCi-20 autoinjector (splitless mode). The GCMS-Solution ver. 1.2 software (Shimadzu Corporation, Kyoto, Japan) was used to interpret chromatographic data. In order to obtain a higher sensitivity of detection, a detector operating in the SIM mode was used to determine the content of individual polycyclic aromatic hydrocarbons. The separation procedures for 25, 27, 44, and 64 characteristic mass ions were tested, and the last method (64 mass ions) was selected for the final PAH analysis.
In order to identify PAHs present in animal bodies, a commercial set of standards for polycyclic aromatic hydrocarbons including the following was used: acenaphthylene, fluorene, phenanthrene, anthracene, pyrene, benzo[a]anthracene, chrysene, benzo[b]fluoranthene, benzo[k]fluoranthene, benzo[a]pyrene, dibenzo[a,h]anthracene, and benzo[g,h,i]perylene. The identification of individual PAHs was carried out based on the retention times of their standards (see Table 1). From a stock solution of 500 µg/mL of individual hydrocarbons, dilutions of standard solutions were prepared in the concentration range of 0.5–50 µg/mL. Then, calibration curves for individual hydrocarbons were prepared based on the calculated area of obtained chromatographic peaks.

2.4. Statistical Analysis

The obtained results were analyzed and checked for normality (Shapiro–Wilk test with Lilliefors correction) and equality of variance (Levene’s test). The significance of differences between the means were tested by one- or two-factor variance analysis (STATISTICA 13.1 software), and the means were differentiated by Fisher’s LSD test at p < 0.05. One-factor variance analysis was performed for PAH content in animals because at some dates, it was not possible to measure this parameter or PAHs were not detected, which make the factor “weeks” impossible to use.

3. Results

3.1. Survival and Body Mass Gain

3.1.1. Lumbricus terrestris L.—Single-Species Experiment

Both contamination and breeding time had a significant effect on the survival rate and body mass gain of L. terrestris. There was also a significant interaction between the two factors (Table S1). Soil contamination with DF caused significant earthworm mortality of nearly 45% and 85% after 3 and 4 weeks of breeding, respectively (Figure 1). A significant negative effect, however weaker than with DF, was also observed with EO. However, this was evident only in the fourth week of the experiment (24% mortality). Mortality in the control did not exceed 10%. Body mass gain, on the other hand, decreased in all treatments (including control) starting from the second week of the experiment (Figure 2). The smallest decrease in body mass was noted in the P treatment (5% vs. 24% in control at week 4 of the experiment).

3.1.2. Lumbricus terrestris L. and Porcellio scaber Latr.—Two-Species Experiment

As in the case of the single-species breeding, in the two-species breeding both culture time and contamination significantly affected the survival rate of earthworms and woodlice (Table S2). In the case of earthworms, the interaction of both factors was also significant. There was a high mortality rate of earthworms from the control group in the combined breeding with woodlice starting from the third week of the experiment (Figure 3). It was higher than in the treatments with P- and EO-contaminated soil. In the fourth week of breeding, significantly lower mortality than in the control was recorded in the soil contaminated with P and DF. In contrast, the survival rate of woodlice in each treatment was similar (Figure 4). Only in the case of EO was it significantly reduced compared to the control at weeks 3 and 4 of breeding.

3.2. TPH Content in Soil

3.2.1. Lumbricus terrestris L.—Single-Species Experiment

Changes in the content of petroleum compounds in the soil contaminated with P in which the single-species test was carried out were insignificant and largely similar to the fluctuations in the TPH content observed in the control soil (Figure 5a, Table S3). The presence of earthworms in the soil contaminated with DF contributed to a decrease in the content of petroleum compounds, mainly during the first two weeks of the experiment, from approx. 7500 to 6000 mg/kg soil d. m. In the second part of the experiment, no further decrease in the amount of TPH in the soil was found, which could be associated with the death of most of the tested animals before the end of the experiment. Still, the differences between separate dates (0, 2, or 4 weeks) were not statistically proven (Figure 5b). On the other hand, during 4 weeks of cultivation of earthworms in soil contaminated with EO, a decrease in the content of petroleum derivatives from approx. 33,000 to approx. 31,000 TPH mg/kg d. m. soil was visible after four weeks. However, the differences between 0 and 4 weeks were statistically insignificant. Significant difference was found only between the second (when TPH content increased) and fourth weeks (Figure 5c).

3.2.2. Lumbricus terrestris L. and Porcellio scaber Latr.—Two-Species Experiment

Figure 6, Figure 7 and Figure 8 present changes in the content of oil derivatives in the soil during the exposure of two-species breeding (L. terrestris + P. scaber) to hydrocarbon pollutants. During the analysis of the content of oil derivatives in the soil, the highest losses of hydrocarbons in the soil were found in the variant of soil contamination with EO (Figure 8). L. terrestris and P. scaber together caused a significant decrease in TPH content in this treatment after 4 weeks (Table S4). The data presented later in the text show that it was in the case of animal exposure to EO that the most significant accumulation of PAHs (especially in the bodies of earthworms) occurred. Therefore, it may mean that these animals contributed to reducing the total petroleum hydrocarbons derived from EO. In the case of P, only time was a significant factor, while in DF treatment, none of the analyzed factors influenced TPH content significantly (Table S4).

3.3. PAHs Concentration in Animals Bodies

3.3.1. Lumbricus terrestris L.—Single-Species Experiment

Table 2 and Table S5 summarizes the detailed results of PAH content in the bodies of earthworms exposed to PPs in a single-species experiment.
Almost all of the analyzed hydrocarbons were identified in the animals’ bodies. The highest amounts of PAHs accumulated in the earthworm’s tissues were found when animals were exposed to EO. In the case of this experimental variant, a tendency was observed to accumulate PAHs (especially anthracene and benzo[b]fluoranthene) in earthworms during the whole experiment. The content of PAHs found in the bodies of earthworms was generally higher than in the two-species experiment (Table 3), but the content of petroleum compounds in the soil used in this variant was also higher. Due to the high mortality rate of earthworms grown in the experimental variant of soil contaminated with diesel oil, the analysis of PAH content could only be performed after two weeks of the experiment.

3.3.2. Lumbricus terrestris L. and Porcellio scaber Latr.—Two-Species Experiment

Table 3 and Table 4 as well as Tables S6 and S7 present detailed results of PAHs content in the bodies of L. terrestris and P. scaber in a two-species experiment. Particularly high concentrations of PAHs were recorded in the case of earthworms. In the body of these animals, all the analyzed hydrocarbons were identified, and their remarkably high values were found in the case of DF and EO exposure. In samples obtained from earthworms exposed to EO high-weight polyaromatic hydrocarbons were observed to accumulate. This is the case of benzo[k]fluoranthene; the content in the body of earthworms after two weeks of the experiment was 0.75 µg/g of the sample, and after another two weeks, it increased to 1.32 µg/g of the sample. In the case of P. scaber tissues, no PAHs were detected in the DF variant, and the highest concentrations of residue hydrocarbons were found in the samples exposed to EO. The higher concentration of hydrocarbons detected in samples derived from earthworms compared to woodlice can probably be explained by the fact that earthworms are much more exposed to direct contact with these xenobiotic substances in the soil due to their lifestyle.

4. Discussion

Our research focused in particular on assessing the possibility of increasing the effectiveness of TPH removal from contaminated soil by using two species from different systematic positions (earthworms and woodlice) simultaneously (obtained data were compared to using earthworms alone). The experiment was conducted in long-term aging soil contamination (i.e., 12 and 24 months after contamination) using petroleum products (which are less frequently analyzed than crude oils). All the aspects mentioned above were examined in terms of their influence on the life parameters of the test animals, the content of TPH in the soil, and the level of PAH accumulation in the bodies of L. terrestris and P. scaber.

4.1. Survival and Body Mass Gain

The survival rate of earthworms in the presence of petroleum contaminants is fairly well recognized. However, it should be noted that this parameter is not only species-specific, but also depends on the type of oil pollutants, their concentration, soil type, climatic conditions, etc. [13,15,28,38,39], which makes comparisons of research results obtained by different authors difficult. In doing so, most of the available results relate to the sensitivity of earthworms to crude oils. L. terrestris, as a deep-burrowing earthworm, is considered more vulnerable to contaminants compared to litter species that feed on the soil surface, such as E. fetida, which is commonly used in toxicological studies. Moreover, L. terrestris does not tend to avoid contact with contaminated soil, even when it has the opportunity to do so [40]. Almutari [14] showed that contamination with Kuwaiti oil at 1.0% caused 60% mortality of L. terrestris after 15 days of the experiment. E. fetida in this study proved to be more resistant in this regard, as a 1.0% contamination concentration caused 30% mortality after 15 days of the experiment. The same concentration (1%) of unweathered Arabian light crude oil caused 75% mortality of L. terrestris after 15 days of exposure, with E. fetida showing lower mortality compared to L. terrestris when the oil was weathered (30%), but higher mortality when the oil was unweathered (90%) [40]. On the other hand, in the case of the tropical earthworm Pontoscolex corethrurus (an endogenous species, like L. terrestris), Cuevas-Diaz et al. [32] found the median lethal concentration (LC50) of TPH in soil contaminated with “Maya” Mexican heavy crude oil to be 3067.32 mg/kg (during a 14-day test). The LC/EC50 for L. terrestris in lubricating oil composed mainly of C10–C16 and >C16–C34 petroleum hydrocarbons was 2158 mg TPH/kg of dry soil (14 days test) [41]. In our single-species experiment, there was no significant increase in L. terrestris mortality under the influence of PPs after 2 weeks, while at a later time, DF and EO caused mortalities of 85% and 24%, respectively, after 4 weeks of exposure to TPH contents in the initial soil of 7445 and 33,141 mg/kg d.w. of soil, respectively. Thus, when compared to the results obtained for crude oils (data mentioned above), the petroleum products in our research seem to be less toxic to L. terrestris. Information on the effect of PPs on earthworm survival is limited. Chachina et al. [42] reported significant mortality of E. fetida earthworms in the presence of DF, reaching 30% after 2 weeks of exposure at contamination levels of 40 g/kg. The earthworm E. eugeniae survived DF concentrations of 5 mL per 1 kg of experimental soil [43]. A study on the toxicity of liquid solutions of various petroleum substances for L. terrestris showed a LC50 for DF of 44.5 µL/L [38], and for P, it was even lower at 37.0 µL/L. In the present experiment, there was no significant effect of P on earthworm mortality. The difference is probably due to the fact that in Ebere’s [44] experiment the PPs were applied at the start of the experiment, while our experiment was conducted 12 months after soil contamination, when the volatiles contained in P had already managed to be released. In turn, the high toxicity of DF in our experiment is noteworthy. Compared to EO, DF contains lighter fractions, which may have been the reason for its higher toxicity to earthworms. Similarly, Martinkosky et al. [13] found significantly higher mortality of E. fetida when exposed to lighter crude oil than to crude oil with high viscosity. However, in a two-species breeding with P. scaber, it was EO that was found to be more toxic to earthworms than DF. This would indicate that the presence of P. scaber promoted the release of substances from EO that had higher toxicity to earthworms. This may be related to the increase in microbial biomass that is observed in the presence of P. scaber [8]. This may also be confirmed by the significant level of TPH reduction in EO-contaminated soil under the activity of earthworms and woodlice in two-species breeding (the highest among the examined PPs) (Figure 6, Figure 7 and Figure 8) as well as the high level of PAHs accumulation in the body of earthworms (Table 3).
In our experiment, only EO caused a significant reduction in woodlice survival after 3 and 4 weeks of exposure during two-species breeding with L. terrestris (to a level of 53% and 36%, respectively). In another experiment on the effect of PPs on P. scaber (single-species breeding), with similar levels of TPH in the initial soil contaminated with EO, there was no significant reduction in the survival rate of P. scaber during 4-week culture [12]. Additionally, other authors confirm the rather high resistance of P. scaber to petroleum products [45,46].
In the available literature, body mass change was much less frequently analyzed than survival rate. Weight gain of earthworm P. corethrurus was a parameter more sensitive to the presence of contaminants than its mortality [32]. After 7 and 14 days of exposure to “Maya” Mexican heavy crude oil (4845 mg TPH/kg), this earthworm responded with decreases in biomass of 29.5% and 35.6%, respectively. In our experiment, the body mass gain of L. terrestris was not significantly affected by the pollutants for a period of 4 weeks of breeding. The differences in the response of different earthworm species in their sensitivity to oil pollution may be related to their morphology, physiology, and lifestyle [39].
Summarizing the data obtained in this experiment in relation to survival and body mass gain, it could be indicated that petroleum products are less toxic than crude oils; there is a reduction in the negative impact of P, resulting from the time elapsed from the moment of contamination; and there is an increase in the toxicity of EO to earthworms in the case of the simultaneous presence of P. scaber.

4.2. TPH Content in Soil

Soil contamination with petroleum compounds causes severe environmental damage by affecting the activity of soil organisms. The efficiency of bioremediation techniques largely depends on the qualitative and quantitative composition of the pollutants released into the natural environment, the type of soil, and the microorganisms present in the contamination area. Earthworms can increase the efficiency of hydrocarbon biotransformation by mixing and aerating contaminated soil, stimulating the initiation of autochthonous microbial biocenoses by secreting surfactants or enzymes into their digestive tract, as well as decomposing toxic compounds using tissue enzymes present in the body of these animals [13]. Schaefer et al. [38] documented a significant loss of TPH during the incubation of soil contaminated with hydrocarbons in the presence of L. terrestris and E. fetida. In the following experimental cycle, the same authors showed the possibility of effective biotransformation of TPH supported by organic additives in soil contaminated with diesel oil (the initial concentration of TPH was 9500 mg/kg d. m.) in the presence of three different species of earthworms (E. fetida, A. chlorotica, and L. terrestris), obtaining in a 28-day experiment the efficiency of oxidation of xenobiotics at the level of 30 to 42% depending on the animal species used [3]. Ameh et al. [47] obtained a 36.28% reduction in TPH in soil with the addition of E. eugeniae earthworms in the test lasting 42 days. However, in the studies conducted by Ekperusi and Aigbodion [31], a very high decrease in the TPH level was obtained (TPH initial concentration in the soil was approx. 700 mg/kg d. m.) during the incubation of soil contaminated with diesel fuel with E. eugeniae earthworms. The loss of TPH reached the levels of 15.2, 67.08, and approx. 85% (after 30, 60, and 90 days of the experiment, respectively). In an experiment carried out with crude-oil-contaminated soil samples with the addition of Hyperiodrilus africanus earthworms, the same authors obtained reductions in TPH level (TPH initial concentration in the soil was approx. 1200 mg/kg d. m.) of 22.01, 44.29, and 68.29% (30, 60, and 90 days, respectively) [31]. During an experiment lasting more than 500 days, Martinkosky et al. [13] investigated the impact of E. fetida earthworms in the bioremediation process of crude-oil-contaminated soil (initial concentration of 30,000 mg/kg d. m.). The observed biotransformation rate was about 90 mg/day, was maintained for 200 days of the experiment, and was then decreased to 20 mg/day until the end of the experiment. On the other hand, Nassar and Said [48] showed the possibility of using Allolobophora caliginosa earthworms in the bioremediation of soil heavily contaminated with light crude oil, achieving a significant hydrocarbon removal efficiency after 60 days of the experiment—from approx. 55 to 71% depending on the initial concentration of xenobiotics used. Our research used soil contaminated with three complex mixtures of hydrocarbons (petrol, diesel fuel, used engine oil). In this experimental setup, we tested the influence of the presence of L. terrestris on the efficacy of the biotransformation/bioremediation process of the applied xenobiotics. During the 4-week experiment in the single-species variant, we obtained results of the reduction in hydrocarbons comparable to those presented by other researchers. The efficiency of the TPH removal in the case of soil contaminated with DF was approximately 21%, and in soil contaminated with EO, it was 9%. Our results indicate that the fractions of hydrocarbons that were more difficult to degrade were hydrocarbons with high molecular weights. However, the highest efficiency of the TPH removal from the soil was observed in the case of soil contaminated with P (approx. 39.7%) the changes in TPH content were statistically insignificant and similar to the fluctuations in the TPH content observed in the control soil.
It should be noted that in the conducted experiments, the biotransformation process is often tested in the presence of a limited number of invertebrates and other organisms, which can affect the progress of this process. However, the natural conditions in which in situ bioremediation processes are often carried out are much more complicated. Therefore, it is necessary to observe the vermiremediation processes in complex systems. The high efficiency of diesel fuel biotransformation (initial content 10,000 µL/kg d. m.) in the presence of E. fetida earthworms is reported in the paper by Fernández et al. [33], in which the experimental microcosm model was used in an attempt to recreate the natural conditions in the soil environment as much as possible. In the case of the most comprehensive experimental variant (earthworms and plant vegetation), the authors obtained reduction in the initial diesel fuel concentration by 41.4 and 72.1% after 90 and 180 days, respectively. Chachina et al. [42] conducted an experiment in which they used E. fetida earthworms to support the bioremediation of soil contaminated with petroleum and diesel oil (initial concentrations of 20,000–60,000 and 20,000–40,000 mg/kg d. m., respectively), inoculating additionally contaminated soil with bacteria of the genus Pseudomonas, Azotobacter, and Clostridium; the yeast Saccharomyces; and the fungi Aspergillus, Penicillium, and Actinomycetales. During the 22-week experiment, a practically complete biodegradation of petroleum hydrocarbons was demonstrated (efficiency at 99% regardless of the initial concentration of hydrocarbons) and about 95% loss of hydrocarbons in the variants of soil contaminated with diesel fuel. In order to simulate a more complex environment of the hydrocarbon degradation process, we tested the bioremediation process assisted by the presence of earthworm L. terrestris and P. scaber isopod species (two-species experimental variant), which to the best of our knowledge has not been examined so far. The efficiencies of TPH biotransformation in soil samples contaminated with DF and EO were 22.8%, and 28.7%, respectively. Our results differ from the previously cited results of experiments of other research groups (e.g., [33] or [42]). However, it is worth noting that the duration of our two-species experimental variant was much shorter (4 weeks) than the experiments of the other two research groups (22 and about 26 weeks, respectively), which—in the case of complex hydrocarbon mixtures—may be of crucial importance. At the same time, our two-species variant was not a typical experimental microcosm model, and the soil used during our research was not additionally inoculated with bacteria of the genus Pseudomonas (which are effective degraders of organic pollutants) or bacteria of the genus Azotobacter belonging to the group of plant-growth-promoting microorganisms. Still, it is worth noticing that in the case of the two-species variant, TPH loss in EO contaminated soil was much higher than in the single-species variant (29% vs. 9%, respectively). This supports our hypothesis that using the two species together can increase the efficiency of the bioremediation process.
Despite the effective biotransformation of hydrocarbons compounds, a significant amount of xenobiotic material remains in the soil after the completion of the experiments. Improving the efficiency of this process may be associated with the extension of the duration of biodegradation experiments, which in laboratory conditions (up to 30 and a maximum of 40 days) differs from the process conditions that often last several months, if not years. Extending the biotransformation process’s time may improve this process’s efficiency, giving the animals time to penetrate the contaminated soil better and increasing the bioavailability of xenobiotics [31,43].

4.3. PAH Concentration in Animals Bodies

In addition to assessing TPH reduction from soil contaminated with oil derivatives, it is essential to analyze the behavior of individual xenobiotics in the organisms of animals exposed to direct contact with hydrocarbons, especially of those compounds belonging to the PAH group.
In their research, Van Straalen and Verweij [49] analyzed the possibility of benzo[a]pyrene bioaccumulation by P. scaber. The animals were supplied with food contaminated with B[a]P, which led to the bioaccumulation of this hydrocarbon in the tissues of P. scaber up to 125 µg/g of tissue. During studies on Lumbricus rubellus Hoff. and P. scaber collected at different distances from the furnace plant, phenanthrene, fluoranthene, pyrene, benzo[a]anthracene, chrysene, benzo[a]pyrene, benzo[b]fluoranthene, and benzo[k]fluoranthene were detected in the tissues of the animals. It was observed that in the tissues of earthworms, these PAHs accumulated in much more significant amounts (1.5 to 4 times more) than in the case of the isopod P. scaber [50] (similarly to in our experiment). An analysis of pyrene biotransformation by P. scaber was carried out by exposing the animals to this xenobiotic introduced into their organisms with contaminated food, as a result of which it was possible to identify 1-hydroxypyrene being the primary metabolite of pyrene biotransformation in P. scaber tissues [51] and other terrestrial invertebrates, e.g., Oniscus asellusi, Eisenia andrei or Folsomia candida [18,19,20]. In our GC-MS analyses of PAH extracts obtained from P. scaber and L. terrestris tissues, it was impossible to identify 1-hydroxypyrene or any other metabolite that could indicate the biotransformation of PAH in the bodies of the tested animals. Zakia et al. [26] compared the bioaccumulation of PAH by two earthworm species E. fetida and L. terrestris (0.204 and 0.084 g/g, respectively). Researchers found only four PAHs in animal tissues: fluoranthene, pyrene, benzo[a]anthracene, and chrysene. There was no accumulation of polycyclic aromatic hydrocarbons with 3, 5, or 6 aromatic rings [26]. Brown et al. [52] investigated the exposure of E. fetida earthworms to phenanthrene (at concentrations of 50–1500 mg/kg d. m. soil). The analysis showed that in the presence of earthworms, the bioavailability of phenanthrene in the soil increased by 65–97% depending on its initial concentration. Although no biodegradation of phenanthrene was found 48 h after the start of the experiment, the analysis of the metabolome showed a response to stress factors through changes in the content of amino acids (isoleucine, alanine, and glutamine) and maltose. Research by MacKelvie et al. [53] in which the biodegradation process of phenanthrene was analyzed (initial concentration approx. 320 mg/kg d.m. soil) showed that the observed 15-day biodegradation of phenanthrene was the result of the activity of autochthonous microbial biocenosis and took place without the participation of E. fetida earthworms. Differently from the group of McKelvie et al., the results were obtained by Contreras-Ramos et al. [54], who in an 11-week experiment showed that the presence of E. fetida earthworms significantly improved the biodegradation efficiency of selected PAHs. The mean PAH biodegradation efficiencies led by autochthonous microorganisms was 23%, 77%, and 13% for anthracene, phenanthrene, and benzo[a]pyrene, respectively. At the same time, the introduction of earthworms into the soil contaminated with these hydrocarbons increased the bioremediation efficiency to 51% and 100% in the cases of anthracene and phenanthrene, respectively. Gomez-Eyles et al. [55] confirmed in their research an increase in the bioavailability (by over 40%) of polycyclic aromatic hydrocarbons in PAH-contaminated soil due to the presence of E. fetida earthworms. Cachada et al. [56] analyzed the possibility of bioaccumulating PAHs in the Eisenia andrei earthworms found in the Lisbon urban soil in Spain. During the research, they observed that PAHs composed of four aromatic rings are the most bioaccumulated compounds of all PAHs identified in tested soil samples, which could be associated with them not being as volatile as 2- and 3-ring hydrocarbons and not as recalcitrant as more complex hydrocarbons. Esmaeili et al. [57] compared the accumulation of 16 US-EPA priority PAH by three ecotypes of earthworms: Amynthas sp., E. fetida, and L. terrestris. The conducted tests showed that Amynthas sp. had the highest level of PAH bioaccumulated (almost 8.7 times more accumulated PAH than E. fetida), while E. fetida had the lowest levels of PAH bioaccumulation.
The results obtained during our experiments, regardless of the variant of the experiment, i.e., single- or two-species experiment, clearly show the tendency to bioaccumulate PAHs in invertebrates’ tissues. The highest increase in PAH content inside animal tissues was recorded in the case of the L. terrestris variant exposed to soil contaminated with EO (single-species experiment). Between the second and the fourth weeks of the experiment, we noted an almost 22-fold increase in the content of PAHs in the tissues of earthworms. Such a significant increase in xenobiotic material amount inside the examined animal tissues may indicate the direction of metabolism towards accumulating toxic compounds and not their expected biotransformation. The content of PAHs found in the bodies of earthworms in the two-species experiment was generally lower, but the content of petroleum compounds in the soil used in this variant was also lower. Previously observed by the group of van Brummelen et al. [50], the relationship between the PAH content in the bodies of P. scaber and L. rubellus was also confirmed in our analysis. Comparing the total PAH content in the bodies of both animals after the completion of the two-species variant of the experiment (4 weeks), we noted nearly 2.7- and 3.7-times higher PAH concentrations (for soil contaminated with P and DF, respectively) in the body of earthworms than in the tissues of P. scaber. Increased PAH content in the bodies of earthworms may be the result of more direct contact with the contaminated soil environment, as a result of which exposure to xenobiotics may occur within the entire surface of the animal body and in the gastrointestinal tract. Another reason for the lower content of polycyclic aromatic hydrocarbons in P. scaber tissues may be the faster elimination rate of PAH compounds in P. scaber. The relatively high rate of PAH (benzo[a]pyrene) biotransformation by P. scaber was noted in the research by Van Brummelen and Van Straalen [58], indicating the breakdown rate of this hydrocarbon at 1.1 µg/day.

5. Conclusions

  • Survival of earthworms cultivated separately (single-species experiment) in soil contaminated with DF 12 months earlier significantly decreased (85% mortality after 4 weeks), while body mass gain was not significantly changed under the influence of tested PPs;
  • Breeding of L. terrestris together with P. scaber contributed to significant mortality of earthworms regardless of treatments, while woodlice showed much greater resistance to PPs (only EO caused a significant reduction in survival of P. scaber after 3 and 4 weeks of exposure to 53% and 36%, respectively);
  • The presence of L. terrestris for 4 weeks in soil contaminated with DF and EO 12 months earlier resulted in reductions in TPH of 21% and 9%, respectively (the differences, however, were not statistically significant). Changes in TPH content in soil contaminated with P were similar to that in control soil;
  • Breeding of L. terrestris together with P. scaber in soil contaminated 24 months earlier with EO resulted in a significant reduction in TPH content after 4 weeks (by 29% compared to the initial soil);
  • During the conducted experiments, the tested animals showed a tendency towards bioaccumulation of polycyclic aromatic hydrocarbons rather than their biodegradation. The content of PAHs in the tissues of L. terrestris was higher than in P. scaber. The highest amounts of PAHs accumulated in the earthworm’s tissues were found when animals were exposed to EO;
  • Use of L. terrestris together with P. scaber can be considered a promising scenario for increasing the effectiveness of remediation of soils contaminated with petroleum products, especially EO. However, further research is necessary to establish conditions preventing excessive mortality of earthworms in such a layout.

Supplementary Materials

The following supporting information can be downloaded at: https://www.mdpi.com/article/10.3390/en15217835/s1, Table S1: Anova result on the survival and body mass gain—single-species experiment; Table S2: Anova result on the survival and body mass gain two-species experiment; Table S3: Anova result on TPH content in soil samples —single-species experiment; Table S4: Anova result on TPH content in soil samples—two-species experiment; Table S5: Anova result on PAHs content in the bodies of Lumbricus terrestris L.—single-species experiment; Table S6: Anova result on PAHs content in the bodies of Lumbricus terrestris L.—two-species experiment; Table S7: Anova result on PAHs content in the bodies of Porcellio scaber Latr.—two-species experiment.

Author Contributions

Conceptualization, J.G.; methodology, J.G. and P.P.; software, J.G. and P.P.; validation, J.G. and I.B.P.; formal analysis, J.G. and I.B.P.; investigation, J.G., P.P. and A.K.; resources, J.G. and P.P.; data curation, J.G. and P.P.; writing—original draft preparation, J.G. and P.P.; writing—review and editing, J.G., P.P. and A.K.; visualization, J.G. and P.P.; supervision, J.G., I.B.P. and P.P.; project administration, J.G., P.P. and A.K.; funding acquisition, J.G., P.P. and A.K. All authors have read and agreed to the published version of the manuscript.

Funding

This research was funded by a subsidy of the Ministry of Education and Science for the University of Agriculture in Krakow for the years 2021–2022.

Data Availability Statement

The data presented in this study are available on request from the corresponding author. The data are not publicly available due to privacy restrictions.

Conflicts of Interest

The authors declare no conflict of interest.

References

  1. Claxton, L.D. The history, genotoxicity, and carcinogenicity of carbon-based fuels and their emissions. Part 3: Diesel and gasoline. Mutat. Res. 2015, 763, 30–85. [Google Scholar] [CrossRef] [PubMed]
  2. Szulińska, E.; Zakrzewski, D.; Kafel, A.; Gospodarek, J.; Rozpędek, K.; Zawisza-Raszka, A. Level of oxidative stress for the land snail Cepaea nemoralis from aged and bioremediated soil contaminated with petroleum products. Environ. Sci Poll. Res. 2022. online ahead of print. [Google Scholar] [CrossRef] [PubMed]
  3. Schaefer, M.; Filser, J. The influence of earthworms and organic additives on the biodegradation of oil contaminated soil. Appl. Soil Ecol. 2007, 36, 53–62. [Google Scholar] [CrossRef]
  4. Tejada, M.; Masciandaro, G. Application of organic waste on a benzo[a]pyrene polluted soil. Response of soil biochemical properties and role of Eisenia fetida. Ecotox. Environ. Safe. 2011, 74, 668–674. [Google Scholar] [CrossRef]
  5. Pasmionka, I.B.; Bulski, K.; Boliglowa, E. The participation of microbiota in the transformation of nitrogen compounds in the soil-a review. Agronomy 2021, 11, 977. [Google Scholar] [CrossRef]
  6. Van Wensem, J.; Verhoef, H.A.; Van Straalen, N.M. Litter degradation stage as a prime factor for isopod interaction with mineralization processes. Soil Biol. Biochem. 1993, 25, 1175–1183. [Google Scholar] [CrossRef]
  7. Zimmer, M.; Topp, W. Microorganisms and cellulose digestion in the gut of the woodlouse Porcellio scaber. J. Chem. Ecol. 1998, 24, 1397–1408. [Google Scholar] [CrossRef]
  8. Kautz, G.; Topp, W. Acquisition of microbial communities and enhanced availability of soil nutrients by the isopod Porcellio scaber (Latr.) (Isopoda: Oniscidea). Biol. Fert. Soils 2000, 31, 102–107. [Google Scholar] [CrossRef]
  9. Zimmer, M. Surfactants in the gut fluids of Porcellio scaber (Isopoda: Oniscidea), and their interactions with phenolics. J. Insect Physiol. 1997, 43, 1009–1014. [Google Scholar] [CrossRef]
  10. Delhoumi, M.; Catania, V.; Zaabar, W.; Tolone, M.; Quatrini, P.; Achouri, M.S. The gut microbiota structure of the terrestrial isopod Porcellionides pruinosus (Isopoda: Oniscidea). Eur. Zool. J. 2020, 87, 357–368. [Google Scholar] [CrossRef]
  11. Zimmer, M.; Kautz, G.; Topp, W. Olfaction in terrestrial isopods (Crustacea: Oniscidea): Responses of Porcellio scaber to the odour of litter. Eur. J. Soil Biol. 1996, 32, 141–147. [Google Scholar]
  12. Gospodarek, J.; Petryszak, P.; Koloczek, H.; Rusin, M. The effect of soil pollution with petroleum-derived substances on Porcellio scaber Latr. (Crustacea, Isopoda). Environ. Monit. Assess. 2019, 191, 38. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  13. Martinkosky, L.; Barkley, J.; Sabadell, G.; Gough, H.; Davidson, S. Earthworms (Eisenia fetida) demonstrate potential for use in soil bioremediation by increasing the degradation rates of heavy crude oil hydrocarbons. Sci. Total Environ. 2017, 580, 734–743. [Google Scholar] [CrossRef] [PubMed]
  14. Almutairi, M. Vermiremediation strategy for remediation of Kuwaiti oil contaminated soil. SN Appl. Sci. 2019, 1, 1312. [Google Scholar] [CrossRef] [Green Version]
  15. Cook, S.V.; Chu, A.; Goodman, R.H. Leachability and toxicity of hydrocarbons, metals and salt contamination from flare pit soil. Wat. Air Soil Poll. 2002, 133, 297–314. [Google Scholar] [CrossRef]
  16. ISO-11268-1:2012; Soil Quality-Effects of Pollutants on Earthworms. Part 1: Determination of Acute Toxicity to Eisenia fetida/Eisenia andrei. International Standard Organization (ISO): Geneva, Switzerland, 2012; 2nd ed.
  17. Calisi, A.; Lionetto, M.G.; De Lorenzis, E.; Leomanni, A.; Schettino, T. Metallothionein induction in the coelomic fluid of the earthworm Lumbricus terrestris following heavy metal exposure: A short report. Biom. Res. Int. 2014. online ahead of print. [Google Scholar] [CrossRef] [Green Version]
  18. Calisi, A.; Zaccarelli, N.; Lionetto, M.G.; Schettino, T. Integrated biomarker analysis in the earthworm Lumbricus terrestris: Application to the monitoring of soil heavy metal pollution. Chemosphere 2013, 90, 2637–2644. [Google Scholar] [CrossRef]
  19. Gospodarek, J. Survival rate of redworms and woodlice in soil contamined with petrol, diesel oil and engine oil. Ecol. Chem. Eng. A 2012, 19, 1369–1374. [Google Scholar] [CrossRef]
  20. Stroomberg, G.J.; Ariese, F.; van Gestel, C.A.M.; van Hattum, N.H.; Velthorst, N.H.; van Straalen, N.M. Pyrene biotransformation products as biomarkers of polycyclic aromatic hydrocarbon exposure in terrestrial Isopoda: Concentration-response relationship, and field study in a contaminated forest. Environ. Toxicol. Chem. 2003, 22, 224–231. [Google Scholar]
  21. Stroomberg, G.J.; Ariese, F.; van Gestel, C.A.M.; van Hattum, N.H.; Velthorst, N.H.; van Straalen, N.M. Pyrene biotransformation and kinetics in the hepatopancreas of the isopod Porcellio scaber. Arch. Environ. Contam. Toxicol. 2004, 47, 324–331. [Google Scholar] [CrossRef]
  22. Stroomberg, G.J.; Zappey, H.; Steen, R.J.C.A.; van Gestel, C.A.M.; Ariese, F.; Velthorst, N.H.; van Straalen, N.M. PAH biotransformation in terrestrial invertebrates—A new phase II metabolite in isopods and springtails. Comp. Biochem. Physiol. C 2004, 138, 129–137. [Google Scholar] [CrossRef] [PubMed]
  23. Luthe, G.; Stroomberg, G.J.; Ariese, F.; Brinkham, U.A.T.; Van Straalen, N.M. Metabolism of 1-fluoropyrene nad pyrene in marine flatfish and terrestrial ispods. Environ. Toxicol. Pharmacol. 2002, 12, 221–229. [Google Scholar] [CrossRef]
  24. Kampe, S.; Schlechtriem, C. Bioaccumulation of hexachlorobenzene in the terrestrial isopod Porcellio scaber. Environ. Toxicol. Chem. 2016, 35, 2867–2873. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  25. Krauss, M.; Wilcke, W.; Zech, W. Availability of polycyclic aromatic hydrocarbons (PAHs) and polychlorinated biphenyls (PCBs) to earthworms in urban soils. Environ. Sci. Technol. 2000, 34, 4335–4340. [Google Scholar] [CrossRef]
  26. Parrish, Z.D.; White, J.C.; Isleyen, M.; Gent, M.P.N.; Iannucci-Berger, W.; Eitzer, B.D.; Kelsey, J.W.; Mattina, M.I. Accumulation of weathered Polycyclic Aromatic Hydrocarbons (PAHs) by plant and earthworm species. Chemosphere 2006, 64, 609–618. [Google Scholar] [CrossRef]
  27. Shelton, M.; Chapman, P.J.; Foss, S.S.; Fisher, W.W. Degradation of weathered oil by mixed marine bacteria and the toxicity of accumulated water-soluble material to two marine Crustacea. Arch. Environ. Con. Tox. 1999, 36, 13–20. [Google Scholar] [CrossRef]
  28. Chaineau, C.; Yepremian, C.; Vidalie, J.; Ducreux, J.; Ballerini, D. Bioremediation of a crude oil-polluted soil: Biodegradation, leaching and toxicity assessments. Water Air Soil Pollut. 2003, 144, 419–440. [Google Scholar] [CrossRef]
  29. Van Hamme, J.D.; Singh, A.; Ward, O.P. Recent advances in petroleum microbiology. Microbiol. Mol. Biol. R. 2003, 67, 503–549. [Google Scholar] [CrossRef] [Green Version]
  30. Njoku, K.L.; Nomba, E.U.; Olatunde, A.M. Vermiremediation of crude oil contaminated soil using Eudrillus euginae and Lumbricus terrestris. J. Biol. Environ. Sci. 2017, 11, 43–50. [Google Scholar]
  31. Ekperusi, O.A.; Aigbodion, F.I. Bioremediation of petroleum hydrocarbons from crude oil-contaminated soil with the earthworm: Hyperiodrilus africanus. 3 Biotech 2015, 5, 957–965. [Google Scholar] [CrossRef] [Green Version]
  32. Cuevas-Diaz, M.C.; Vazquez-Luna, D.; Martinez-Hernandez, S.; Guzman-Lopez, O.; Ortiz-Ceballos, A.I. Sensitivity of the endogeic tropical earthworm Pontoscolex corethrurus to the presence of heavy crude oil. Bull. Environ. Contam. Toxicol. 2017, 99, 154–160. [Google Scholar] [CrossRef] [PubMed]
  33. Fernandez, M.D.; Pro, J.; Alonso, C.; Aragonese, P.; Tarazona, J.V. Terrestrial microcosms in a feasibility study on the remediation of diesel-contaminated soils. Ecotox. Environ. Safe. 2011, 74, 2133–2140. [Google Scholar] [CrossRef] [PubMed]
  34. Wen, J.B.; Zhang, J.J.; Wang, Z.H.; Zhang, Y.X. Correlations between emulsification behaviors of crude oil-water systems and crude oil compositions. J. Petrol. Sci. Eng. 2016, 146, 1–9. [Google Scholar] [CrossRef]
  35. Michel, J.; Fingas, M. Oil Spills: Causes, consequences, prevention, and countermeasures. In Fossil Fuels: Current Status and Future Directions; World Scientific Publishing Company: Hackensack, NJ, USA, 2016; pp. 159–201. ISBN 9789814699983. [Google Scholar]
  36. Gospodarek, J.; Petryszak, P.; Koloczek, H. The effect of the bioremediation of soil contaminated with petroleum derivatives on the occurrence of epigeic and edaphic fauna. Biorem. J. 2016, 20, 38–53. [Google Scholar] [CrossRef]
  37. PN-C-04573-01:1986; Determination of the Total Content of Organic Substances Extractable with Petroleum Ether by Weight Method. Polish Standardization Committee: Warsaw, Poland, 1986. (In Polish)
  38. Schaefer, M.; Petersen, S.O.; Filser, J. Effects of Lumbricus terrestris, Allolobophora chlorotica and Eisenia fetida on microbial community dynamics in oil-contaminated soil. Soil Biol. Biochem. 2005, 37, 2065–2076. [Google Scholar] [CrossRef]
  39. Lowe, C.N.; Butt, K.R. Earthworm culture, maintenance and species selection in chronic ecotoxicological studies: A critical review. Eur. J. Soil Biol. 2007, 43, 281–288. [Google Scholar] [CrossRef]
  40. Hanna, S.H.S.; Weaver, R.W. Earthworm survival in oil contaminated soil. Plant Soil 2002, 240, 127–132. [Google Scholar] [CrossRef]
  41. Gainer, A.; Cousins, M.; Hogan, N.; Siciliano, S.D. Petroleum hydrocarbon mixture toxicity and a trait-based approach to soil invertebrate species for site-specific risk assessments. Environ. Toxicol. Chem. 2018, 37, 2222–2234. [Google Scholar] [CrossRef]
  42. Chachina, S.B.; Voronkova, N.A.; Baklanova, O.N. Biological remediation of the petroleum and diesel contaminated soil with earthworms Eisenia fetida. Oil Gas Eng. 2016, 152, 122–133. [Google Scholar] [CrossRef] [Green Version]
  43. Ekperusi, O.A.; Aigbodion, I.F. Bioremediation of heavy metals and petroleum hydrocarbons in diesel contaminated soil with the earthworm: Eudrilus eugeniae. SpringerPlus 2015, 4, 540. [Google Scholar] [CrossRef] [Green Version]
  44. Ebere, A.G.; Akintonwa, A. Acute toxicity studies with earthworms, Lumbricus terrestris. Bull. Environ. Contam. Toxicol. 1995, 55, 766–770. [Google Scholar] [CrossRef] [PubMed]
  45. Van Brummelen, T.C.; Stuijfzand, S.C. Effects of benzo[a]pyrene on survival, growth, and energy reserves in the terrestrial isopods Oniscus asellus and Porcellio scaber. Sci. Total Environ. 1993, 134 (Suppl. S2), 921–930. [Google Scholar] [CrossRef]
  46. Faulkner, B.C.; Lochmiller, R.L. Increased abundance of terrestrial isopod populations in terrestrial ecosystems contaminated with petrochemical wastes. Arch. Environ. Contam. Toxicol. 2000, 39, 86–90. [Google Scholar] [CrossRef]
  47. Ameh, A.O.; Mohammed-Dabo, I.A.; Ibrahim, S.; Ameh, J.B.; Tanimu, Y.; Bello, T.K. Effect of earthworm inoculation on the bioremediation of used engine oil contaminated soil. Int. J. Biol. Chem. Sci. 2012, 6, 493–503. [Google Scholar] [CrossRef] [Green Version]
  48. Nassar, S.E.; Said, R.M. Bioremediation assessment, hematological, and biochemical responses of the earthworm (Allolobophora caliginosa) in soil contaminated with crude oil. Environ. Sci. Pollut. Res. 2021, 28, 54565–54574. [Google Scholar] [CrossRef] [PubMed]
  49. Van Straalen, N.M.; Verweij, R.A. Effects of benzo[a]pyrene on food assimilation and growth efficiency in Porcellio scaber (Isopoda). Bull. Environ. Contam. Toxicol. 1991, 46, 134–140. [Google Scholar] [CrossRef]
  50. Van Brummelen, T.C.; Verweij, R.A.; Wedzinga, S.A.; Van Gestel, C.A.M. Polycyclic aromatic hydrocarbons in earthworms and isopods from contaminated forest soils. Chemosphere 1996, 32, 315–341. [Google Scholar] [CrossRef]
  51. Stroomberg, G.J.; Reuther, C.; Kozin, I.; Van Brummelen, T.C.; Van Gestel, C.A.M.; Gooijer, C.; Cofino, W.P. Formation of pyrene metabolites by the terrestrial isopod Porcellio scaber. Chemosphere 1996, 33, 1905–1914. [Google Scholar] [CrossRef]
  52. Brown, S.A.; McKelvie, J.R.; Simpson, A.J.; Simpson, M.J. 1H NMR metabolomics of earthworm exposure to sub-lethal concentrations of phenanthrene in soil. Environ. Pollut. 2010, 158, 2117–2123. [Google Scholar] [CrossRef]
  53. McKelvie, J.R.; Wolfe, D.M.; Celejewski, M.; Simpson, A.J.; Simpson, M.J. Correlations of Eisenia fetida metabolic responses to extractable phenanthrene concentrations through time. Environ. Pollut. 2010, 158, 2150–2157. [Google Scholar] [CrossRef]
  54. Contreras-Ramos, S.M.; Alvarez-Bernal, D.; Dendooven, L. Eisenia fetida increased removal of polycyclic aromatic hydrocarbons from soil. Environ. Pollut. 2006, 141, 396–401. [Google Scholar] [CrossRef] [PubMed]
  55. Gomez-Eyles, J.L.; Sizmur, T.; Collins, C.D.; Hodson, M.E. Effects of biochar and the earthworm Eisenia fetida on the bioavailability of polycyclic aromatic hydrocarbons and potentially toxic elements. Environ. Pollut. 2011, 159, 616–622. [Google Scholar] [CrossRef] [PubMed]
  56. Cachada, A.; Coelho, C.; Gavina, A.; Dias, A.C.; Patinha, C.; Reis, A.P.; Ferreira da Silva, E.; Duarte, A.C.; Pereira, R. Availability of polycyclic aromatic hydrocarbons to earthworms in urban soils and its implications for risk assessment. Chemosphere 2018, 191, 196–203. [Google Scholar] [CrossRef] [PubMed]
  57. Esmaeili, A.; Knox, O.; Juhasz, A.; Wilson, S.C. Differential accumulation of polycyclic aromatic hydrocarbons (PAHs) in three earthworm ecotypes: Implications for exposure assessment on historically contaminated soils. Environ. Adv. 2022, 7, 100175. [Google Scholar] [CrossRef]
  58. Van Brummelen, T.C.; Van Straalen, N.M. Uptake and elimination of benzo[a]pyrene in the terrestrial isopod Porcellio scaber. Arch. Environ. Contam. Toxicol. 1996, 31, 277–285. [Google Scholar] [CrossRef]
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Figure 1. Lumbricus terrestris L. survival—single-species experiment, P—petrol-contaminated soil, EO—engine-oil-contaminated soil, DF—diesel-fuel-contaminated soil. Means marked with the same letters do not differ significantly according to an LSD test at p < 0.05. Factors: weeks × contamination. Vertical bars represent 0.95 confidence intervals.
Figure 1. Lumbricus terrestris L. survival—single-species experiment, P—petrol-contaminated soil, EO—engine-oil-contaminated soil, DF—diesel-fuel-contaminated soil. Means marked with the same letters do not differ significantly according to an LSD test at p < 0.05. Factors: weeks × contamination. Vertical bars represent 0.95 confidence intervals.
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Figure 2. Lumbricus terrestris L. body mass gain—single-species experiment. For explanations, see Figure 1. Means marked with the same letters do not differ significantly according to an LSD test at p < 0.05. Factors: weeks × contamination. Vertical bars represent 0.95 confidence intervals.
Figure 2. Lumbricus terrestris L. body mass gain—single-species experiment. For explanations, see Figure 1. Means marked with the same letters do not differ significantly according to an LSD test at p < 0.05. Factors: weeks × contamination. Vertical bars represent 0.95 confidence intervals.
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Figure 3. Lumbricus terrestris L. survival—two-species experiment. For explanations, see Figure 1. Means marked with the same letters do not differ significantly according to an LSD test at p < 0.05. Factors: weeks × contamination. Vertical bars represent 0.95 confidence intervals.
Figure 3. Lumbricus terrestris L. survival—two-species experiment. For explanations, see Figure 1. Means marked with the same letters do not differ significantly according to an LSD test at p < 0.05. Factors: weeks × contamination. Vertical bars represent 0.95 confidence intervals.
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Figure 4. Porcellio scaber Latr. survival—two-species experiment. For explanations, see Figure 1. Means marked with the same letters do not differ significantly according to an LSD test at p < 0.05. Factors: weeks × contamination. Vertical bars represent 0.95 confidence intervals.
Figure 4. Porcellio scaber Latr. survival—two-species experiment. For explanations, see Figure 1. Means marked with the same letters do not differ significantly according to an LSD test at p < 0.05. Factors: weeks × contamination. Vertical bars represent 0.95 confidence intervals.
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Figure 5. Comparison of changes in TPH content in soil samples contaminated with (a) petrol (P); (b) diesel fuel (DF); and (c) engine oil (EO) during the biodegradation process involving Lumbricus terrestris L—single-species experiment. Means marked with the same letters do not differ significantly according to an LSD test at p < 0.05. Factors: weeks × contamination. Vertical bars mean ± SE.
Figure 5. Comparison of changes in TPH content in soil samples contaminated with (a) petrol (P); (b) diesel fuel (DF); and (c) engine oil (EO) during the biodegradation process involving Lumbricus terrestris L—single-species experiment. Means marked with the same letters do not differ significantly according to an LSD test at p < 0.05. Factors: weeks × contamination. Vertical bars mean ± SE.
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Figure 6. Change in TPH content in soil samples contaminated with petrol during the biodegradation process—two-species experiment. Control—uncontaminated soil without animals; control + animals—uncontaminated soil with animals; P—petrol-contaminated soil without animals; P + animals—petrol-contaminated soil with animals. Means marked with the same letters do not differ significantly according to an LSD test at p < 0.05. Factors: weeks × contamination. Vertical bars mean ± SE.
Figure 6. Change in TPH content in soil samples contaminated with petrol during the biodegradation process—two-species experiment. Control—uncontaminated soil without animals; control + animals—uncontaminated soil with animals; P—petrol-contaminated soil without animals; P + animals—petrol-contaminated soil with animals. Means marked with the same letters do not differ significantly according to an LSD test at p < 0.05. Factors: weeks × contamination. Vertical bars mean ± SE.
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Figure 7. Change in TPH content in soil samples contaminated with diesel fuel during the biodegradation process—two-species experiment. Control—uncontaminated soil without animals; Control + animals—uncontaminated soil with animals; DF—diesel-fuel-contaminated soil without animals; DF + animals—diesel-fuel-contaminated soil with animals. Means marked with the same letters do not differ significantly according to an LSD test at p < 0.05. Factors: weeks × contamination. Vertical bars mean ± SE.
Figure 7. Change in TPH content in soil samples contaminated with diesel fuel during the biodegradation process—two-species experiment. Control—uncontaminated soil without animals; Control + animals—uncontaminated soil with animals; DF—diesel-fuel-contaminated soil without animals; DF + animals—diesel-fuel-contaminated soil with animals. Means marked with the same letters do not differ significantly according to an LSD test at p < 0.05. Factors: weeks × contamination. Vertical bars mean ± SE.
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Figure 8. Change of TPH content in soil samples contaminated with engine oil during the biodegradation process—two-species experiment. Control—uncontaminated soil without animals; Control + animals—uncontaminated soil with animals; EO—engine-oil-contaminated soil without animals; EO + animals—engine-oil-contaminated soil with animals. Means marked with the same letters do not differ significantly according to an LSD test at p < 0.05. Factors: weeks × contamination. Vertical bars mean ± SE.
Figure 8. Change of TPH content in soil samples contaminated with engine oil during the biodegradation process—two-species experiment. Control—uncontaminated soil without animals; Control + animals—uncontaminated soil with animals; EO—engine-oil-contaminated soil without animals; EO + animals—engine-oil-contaminated soil with animals. Means marked with the same letters do not differ significantly according to an LSD test at p < 0.05. Factors: weeks × contamination. Vertical bars mean ± SE.
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Table
Table 1. Retention times and selected ions identified for individual polycyclic aromatic hydrocarbons.
Table 1. Retention times and selected ions identified for individual polycyclic aromatic hydrocarbons.
Compound NameTR [min]Selected Ions Monitoring [m/z]
acenaphthylene31.88263, 75, 76, 126, 150, 151, 152, 153, 154,
fluorene35.25363, 82, 139, 163, 164, 165, 166, 167,
phenanthrene40.02076, 88, 89, 150, 151, 152, 176, 177, 178, 179,
anthracene40.28276, 88, 89, 150, 151, 152, 176, 177, 178, 179,
pyrene47.24488, 100, 101, 198, 199, 200, 201, 202, 203,
benzo[a]anthracene53.418100, 101, 112, 113, 114, 224, 226, 227, 228, 229,
chrysene53.638114, 200, 202, 224, 225, 226, 227, 228, 229,
benzo[b]fluoranthene58.699113, 124, 125, 126, 213, 248, 250, 251, 252, 253,
benzo[k]fluoranthene58.867112, 113, 124, 125, 126, 248, 250, 251, 252, 253,
benzo[a]pyrene60.306112, 113, 124, 125, 126, 248, 250, 251, 252, 253,
dibenzo[ah]anthracene66.100125, 137, 138, 139, 274, 276, 277, 278, 279,
benzo[ghi]perylene67.645124, 125, 136, 137, 138, 274, 275, 276, 277,
1-methylchryseneIS119, 120, 121, 226, 239, 240, 241, 242, 243,
Table
Table 2. Changes in PAH content [µg/g of sample] in the bodies of Lumbricus terrestris L. exposed to petroleum-derived pollutants during the 4-week-long single-species variant after two and four weeks of the experiment. For explanations, see Figure 1; nd—not detected.
Table 2. Changes in PAH content [µg/g of sample] in the bodies of Lumbricus terrestris L. exposed to petroleum-derived pollutants during the 4-week-long single-species variant after two and four weeks of the experiment. For explanations, see Figure 1; nd—not detected.
PDFEO
242424
acenaphthylenend0.01 ± 0.01 a *nd-nd0.05 ± 0.01 a
fluorene0.10 ± 0.02 a0.22 ± 0.01 a0.44 ± 0.02 b-0.04 ± 0.01 a1.27 ± 0.11 c
phenanthrene0.06 ± 0.01 and0.16 ± 0.01 a-0.16 ± 0.01 a1.11 ± 0.09 b
anthracene0.06 ± 0.01 a0.07 ± 0.01 a0.26 ± 0.02 b-0.11 ± 0.01 a2.17 ± 0.07 c
pyrene0.01 ± 0.01 a0.11 ± 0.01 bnd-0.01 ± 0.01 a0.20 ± 0.01 c
benzo[a]anthracene0.06 ± 0.01 bc0.07 ± 0.01 c0.01 ± 0.01 a-0.04 ± 0.01 ab0.12 ± 0.01 d
chrysenend0.06 ± 0.01 and-nd0.10 ± 0.01 a
benzo[b]fluoranthenend1.26 ± 0.08 and-nd5.10 ± 0.21 b
benzo[k]fluoranthene0.66 ± 0.05 c0.61 ± 0.02 c0.42 ± 0.03 b-0.11 ± 0.01 and
benzo[a]pyrenend0.35 ± 0.02 bnd-nd0.12 ± 0.01 a
dibenzo[ah]anthracenendndnd-ndnd
benzo[ghi]perylenend0.05 ± 0.01 and-0.01 ± 0.01 a0.23 ± 0.01 b
* Means ± SE in lines marked with the same letters do not differ significantly according to an LSD test at p < 0.05.
Table
Table 3. Changes in PAH content [µg/g of sample] in the bodies of Lumbricus terrestris L. exposed to petroleum-derived pollutants during the 4-week-long two-species variant after two and four weeks of experimentation. For explanations see Figure 1; nd—not detected.
Table 3. Changes in PAH content [µg/g of sample] in the bodies of Lumbricus terrestris L. exposed to petroleum-derived pollutants during the 4-week-long two-species variant after two and four weeks of experimentation. For explanations see Figure 1; nd—not detected.
PDFEO
242424
acenaphthylene0.03 ± 0.01 a *0.01 ± 0.01 and0.22 ± 0.01 c0.09 ± 0.01 b0.21 ± 0.01 c
fluorene0.07 ± 0.01 a0.04 ± 0.01 a0.09 ± 0.01 a0.35 ± 0.03 c0.20 ± 0.02 b0.22 ± 0.02 b
phenanthrene0.07 ± 0.01 a0.04 ± 0.01 and0.05 ± 0.01 a0.12 ± 0.01 b0.29 ± 0.01 c
anthracene0.10 ± 0.01 ab0.06 ± 0.01 a0.09 ± 0.01 ab0.23 ± 0.02 c0.12 ± 0.01 b0.27 ± 0.02 d
pyrene0.03 ± 0.01 a0.02 ± 0.01 a0.04 ± 0.01 ab0.08 ± 0.01 c0.07 ± 0.01 bc0.15 ± 0.01 d
benzo[a]anthracene0.04 ± 0.01 a0.04 ± 0.01 a0.03 ± 0.01 a0.04 ± 0.01 a0.06 ± 0.01 a0.10 ± 0.01 a
chrysene0.01 ± 0.01 a0.03 ± 0.01 and0.03 ± 0.01 a0.02 ± 0.01 a0.10 ± 0.01 a
benzo[b]fluoranthenend0.07 ± 0.01 and0.48 ± 0.03 bnd0.15 ± 0.02 a
benzo[k]fluoranthene0.70 ± 0.03 cd0.54 ± 0.04 b0.21 ± 0.02 a0.60 ± 0.04 bc0.75 ± 0.05 d1.32 ± 0.10 a
benzo[a]pyrene0.12 ± 0.01 a0.18 ± 0.01 a0.24 ± 0.02 a0.55 ± 0.03 cnd0.80 ± 0.06 d
dibenzo[ah]anthracenend0.01 ± 0.01 and0.03 ± 0.01 a0.01 ± 0.01 a0.04 ± 0.01 a
benzo[ghi]perylene0.18 ± 0.02 c0.10 ± 0.01 b0.07 ± 0.01 ab0.04 ± 0.01 a0.04 ± 0.01 a0.16 ± 0.01 c
* Means ± SE in lines marked with the same letters do not differ significantly according to an LSD test at p < 0.05.
Table
Table 4. Changes in PAH content [µg/g of sample] in the bodies of Porcellio scaber Latr. exposed to petroleum-derived pollutants during the 4-week-long two-species variant after four weeks of experimentation. For explanations see Figure 1; nd—not detected.
Table 4. Changes in PAH content [µg/g of sample] in the bodies of Porcellio scaber Latr. exposed to petroleum-derived pollutants during the 4-week-long two-species variant after four weeks of experimentation. For explanations see Figure 1; nd—not detected.
PDFEO
acenaphthylenendndnd
fluorene0.030 ± 0.01ndnd
phenanthrenendndnd
anthracene0.281 ± 0.02ndnd
pyrenendnd0.562 ± 0.01
benzo[a]anthracenendnd0.177 ± 0.02
chrysene0.027 ± 0.01 a *nd0.208 ± 0.01 a
benzo[b]fluoranthenendndnd
benzo[k]fluoranthenendndnd
benzo[a]pyrenendndnd
dibenzo[ah]anthracene0.085 ± 0.02 and0.072 ± 0.01 a
benzo[ghi]perylenendndnd
* Means ±SE in lines marked with the same letters do not differ significantly according to an LSD test at p < 0.05.
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Gospodarek, J.; Petryszak, P.; Kafel, A.; Paśmionka, I.B. Porcellio scaber Latr. and Lumbricus terrestris L.—PAHs Content and Remediation of Long-Term Aging Soil Contamination with Petroleum Products during a Single- and Two-Species Experiment. Energies 2022, 15, 7835. https://doi.org/10.3390/en15217835

AMA Style

Gospodarek J, Petryszak P, Kafel A, Paśmionka IB. Porcellio scaber Latr. and Lumbricus terrestris L.—PAHs Content and Remediation of Long-Term Aging Soil Contamination with Petroleum Products during a Single- and Two-Species Experiment. Energies. 2022; 15(21):7835. https://doi.org/10.3390/en15217835

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Gospodarek, Janina, Przemysław Petryszak, Alina Kafel, and Iwona B. Paśmionka. 2022. "Porcellio scaber Latr. and Lumbricus terrestris L.—PAHs Content and Remediation of Long-Term Aging Soil Contamination with Petroleum Products during a Single- and Two-Species Experiment" Energies 15, no. 21: 7835. https://doi.org/10.3390/en15217835

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