Representativeness of Folsomia candida, Folsomia fimetaria and Hypoaspis aculeifer for in-soil

For in-soil arthropod invertebrates it was evaluated whether F. candida/F.fimetaria and H. aculeifer are suitable species to represent in-soil arthropod invertebrates (i.e. other springtails and mites other groups that are not tested, e.g. macroarthropods) in thefirst-tier risk assessment.

F. candida and F. fimetaria compared with other springtails

No systematic studies have been published on the sensitivity of F. candida and/or F. fimetaria compared with other collembolans. Table 34 shows three studies from the public literature. From these, with the exception of the experiments with picoxystrobin in Schnug et al. (2014), it seems that tests on F. candida or F. fimetaria with an appropriate assessment factor could be representative of other Collembola. According to the Schnug et al. (2014), the absence of effects on F. fimetaria might be due to food and habitat preferences, causing a much lower exposure. However, it is uncertain whether these effects would be also absent in a standard laboratory toxicity test.

In conclusion, very little data are available to compare the sensitivity of F. candida and F.fimetaria with other springtails. The few available data indicate that F. candida or F. fimetaria could be representative of other springtails with regard to their toxicological sensitivity and it is therefore recommended to use it as a surrogate test species for Collembola considering an appropriate assessment factor. In one case, F. fimetaria appeared to be insensitive compared with other species, but this might be a consequence of food and habitat preferences combined with the test conditions.

H. aculeifer compared with other mites

Lokke and Van Gestel (1998) compared the sensitivity of the herbivorous, oribatid mite Platynothrus peltifer with the predatory mite H. aculeifer for copper chloride, LAS and dimethoate. The results show that EC₅₀ for reproduction does not differ by more than a factor of 2. For Oppia nitens, test methods were also available (Princz et al., 2010), however, no data were found that allow comparison of the sensitivity of this species with other mite species.

Since hardly any data were found comparing the sensitivity of Hypoaspis aculeifer with other soil mites, no conclusion can be drawn concerning the representativeness and further research is required on this issue.

F. candida compared with Hypoaspis aculeifer

From the EFSA conclusions on active substances, chronic toxicity data on Folsomia candida and Hypoaspis aculeifer were extracted. For completeness, sub-lethal toxicity data on Eisenia sp. were also extracted.

For 51 cases and 30 PPPs (10 herbicides, 11 fungicides, 8 insecticides, one acaricide), long-term toxicity data both on Folsomia candida and Hypoaspis aculeifer were available (see Figure 24).

Hypoaspis was more sensitive than Folsomia for 8/51 compounds (15.7%). However, for three cases out of those eight, Eisenia was the most sensitive in-soil organism among the species tested.

Hypoaspis showed a similar sensitivity than Folsomia for 22 compounds out of 51 (41%). These results Table 33: Overview of tests with springtails, comparing toxicity between species

Reference Test design Results for tested organic pesticides

Wiles and Frampton (1996)

Bioassays,field contaminated soil was studied in the lab. Test was

conducted on four species

Residues of cypermethrin and pirimicarb were of low toxicity, causing less than 10% mortality; residues of chlorpyrifos were toxic to all four species of Collembolan (from most to least susceptible)

S. viridis> F. candida > I. palustris > I. viridis

Schnug et al. triclosan in a soil multispecies test system

For esfenvalerate differences in LC₅₀between species were within factor of 2. All species were not sensitive for triclosan, for picoxystrobin large differences in sensitivity (5 orders of magnitude between LC₅₀of most

(P.firmata) and less sensitive species (F. fimetaria)). This difference might be due to food and habitat preferences Lokke and Van

Direct comparison was difficult because endpoints and soils are different between tested in-soil organisms.

Generally, no systematic differences were found between the three species

were confirmed by Owojori et al. (2014), who reported lower to intermediate sensitivity of H. aculeifer in comparison with other in-soil organisms for which standardised tests are available (E. fetida, F. candida, and Enchytraeus albidus). Huguier et al. (2015) also concluded that, compared with other in-soil meso-fauna invertebrates, mites were in general as sensitive or less sensitive than other test species, depending on the studied endpoints and chemicals. The status of H. aculeifer as the only predator among organisms for which a standardised protocol is available, however, highlights its usefulness in ecotoxicological laboratory studies. Please see Appendix H for more information about the data used in the analysis. In the standard laboratory test, however, H. aculeifer is not exposed via the route food uptake since it is fed with clean prey organisms. For exposure via soil, H. aculeifer does not always add information concerning the toxicity of in-soil organisms to tested chemicals compared with the test with F. candida. It is therefore recommended to study the possibility of adapting the test guideline in order to take the food-uptake route into account, which would be relevant for a predator such as H. aculeifer. Further research is needed in order to assess the sensitivity of H. aculeifer since it is not clear whether H. aculeifer is representative of other soil mites. Since F. candida is the most sensitive species in most of the cases, the mode of action of the substance does not seem to be determinant of the response when comparing F. candida, H. aculeifer and E. fetida.

In conclusion, it was found that H. aculeifer is the most sensitive test species for only 3/51 cases.

This result indicates that it might be sufficient to test E. fetida and F. candida with the present test design. H. aculeifer represents another trophic level (predator species) in the standard test battery.

Since it remains unclear if H. aculeifer itself is a good representative for soil mites and the feed-uptake route is not included, it is recommended to adapt the test protocol accordingly.

Folsomia candida compared with Isopods

For the comparison of the sensitivity of F. candida with Isopods, a review study was done based on literature (ISI WoK) and database search (US-EPA, PPDB database, PAN pesticide database).

Toxicity data on Isopods and Collembola were found for 13 compounds (see Table35 and Table 36). Most of the toxicity data reported for isopods were based on mortality or parameters related to individual growth and feeding performance. Data reporting effects on reproduction and avoidance behaviour are scarce. The best represented isopod species is Porcellionides pruinosus, a species with a widespread distribution, being very abundant in southern European and tropical countries. This is considered an‘in-soil’ species, since it can be found in the soil and not predominantly in litter, unlike most isopod species. It is also abundant near agricultural areas. For Collembola, toxicity data found were mostly on mortality and reproduction, with few references on avoidance behaviour. As expected, Folsomia candida was the best represented species.

The analysis of the toxicity data for Isopoda was done first by comparing the toxicity considering exposure via food and soil. Data for the different isopod species were pooled and differences between soil types were taken into account when possible. When more than one value exists for the same parameter and same exposure pathway, the geometric mean was calculated.

0.01 0.1 1 10 100 1000

0.01 0.1 1 10 100 1000

Folsomia28-day NOEC(mg/kg soil)

Hyapoaspis 14-day NOEC (mg/kg soil)

Folsomia vs.Hyapoaspis

Figure 24: Comparison of sensitivity to a set of PPPs (active substances, metabolites or formulations) of Folsomia candida and Hypoaspis aculeifer. The solid line indicates a 1:1 relationship between Folsomia candida and Hypoaspis aculeifer. The light grey area in the figure indicates where Folsomia is more sensitive to pesticides

Table 34: Number of bibliographic records found per isopods and collembolan species and different PPPs Circoniscus

ornatus

Porcellio dilatatus

Porcellio scaber

Porcellionides pruinosus

Folsomia candida

Folsomia fimetaria

Orchesella cincta

Sinella curviseta

Abamectin 5 4 2

Atrazine 1 2 2

Benomyl 8 10

Carbofuran 3 4

Copper 2 1 1

Diazinon 14 6 4

Dimethoate 1 8 6 14

Endosulfan 1 3

Glyphosate 1 3

Imidacloprid 9 4

L-cyhalothrin 5 18 3 1

Lindane 5 6

Spirodiclofen 1 1

Table 35: Acute and chronic endpoints measured in Isopoda and Folsomia candida for several PPPs

Isopods (food) Isopods (soil) Folsomia spp.

Abamectin LC₅₀(Nat soil) 71 LC₅₀(Nat soil) 5.1

NOEC_growth (Nat soil) 3 EC50(Nat soil) 1.4

Atrazine AC₅₀* (Nat soil) 153.1 NOEC_repro 40

LOEC_repro 80

EC50(Nat soil) 43.2

Benomyl LC₅₀ 34,679.7 LC₅₀(Sandy soil) 1,068.8 NOEC_mort (Nat soil) 1

NOEC_growth 1 LC₅₀(OECD) 13–18

EC50(OECD) 8–11

EC₅₀(Nat soils) 2–10

AC₅₀(OECD) 4–15

Carbofuran LC50 486 LC50(Sandy soil) 31.02 NOEC_repro (OECD) 0.01

LC₅₀(Nat soils) 0.06–0.09 EC50(Nat soils) 0.06–0.12

Copper AC₅₀(Nat soil) 922.1 AC₅₀(OECD) 18

EC₅₀(Nat soil) 1200

Diazinon LC50 182.6 LC50(Sandy soil) 3.34 LC50(OECD) 0.1

NOEC_growth > 100 NOEC_repro (OECD) 0.01

NOEC_feeding > 100 EC₅₀(Nat soil) 0.288

Dimethoate LC50 > 75 LC50(Nat soil) 34 LC50(Nat soil) 1.5

LC₅₀(OECD) 44 LC₅₀(OECD) 0.6

EC₅₀_growth (Nat soil) 17.5 NOEC_repro (Nat soil) 2.7 EC50_growth (OECD) 41.2 EC50_repro (Nat soil) 0.77

AC₅₀(Nat soil) 33.2 NOEC_repro (OECD) 2.7

Endosulfan NOEC_feeding 50 LC₅₀(Nat soil) 0.08

AC50(Nat soil) 0.5

EC₅₀(Nat soil) 0.05

Glyphosate AC₅₀(Nat soil) 39.7 EC₅₀_repro (Nat soil) 0.42

NOEC_avoid (Nat soil) 1.2

Imidacloprid NOEC_mortality > 25 LC₅₀(TAS) 20.96

NOEC _growth > 25 NOEC_mort (TAS) 10

NOEC _feeding 10 EC50(TAS) 0.06

NOEC (TAS) 0.01

L-cyhalothrin NOEC_mort (TAS**&OECD) 0.16 NOEC_repro (OECD) 7.6

NOEC_mort (Nat soils) 0.17 EC50(OECD 5%) 5.64

LC₅₀(TAS&OECD) 0.61

LC₅₀(Nat soils) 0.31

NOEC_repro (OECD) 0.1

NOEC_repro (Nat soil) 0.1

EC₅₀_repro (OECD) 0.4

EC50_repro (Nat soil) 0.13

Lindane LC₅₀(OECD) 80 LC₅₀ 1

AC₅₀(Nat soil) 35.3 EC₅₀(OECD) 0.13

NOEC (OECD) 0.03

EC₅₀(Nat soil) 0.8

Spirodiclofen AC₅₀(Nat soil) 0.9 EC₅₀_repro (Nat soil) 0.65

LC50: lethal concentration, median; NOEC: no observed effect concentration.

*: AC50: concentration inducing avoidance in 50% of the tested animals

**: TAS: Tropical artificial soil.

For the four compounds where comparable data are available (mortality data only), contact exposure to soil was revealed to lead to more adverse effects for isopods than food exposure.

Although data are scarce, toxicokinetic studies performed by Sousa et al. (2000) with lindane revealed that isopods were able to accumulate more when exposed via soil than when exposed via food (the reason was mainly the high excretion rates observed and the slower degradation of the compound in the soil matrix). Despite this trend, this comparison between exposure routes should be interpreted carefully since none of these studies considered another important exposure route for these animals, contact in litter.

The sensitivity of Isopoda and Collembola was compared for soil exposure only using the same parameters and measurement endpoints, directly for eight compounds (mainly acute data) and indirectly for five of the compounds. In this last comparison, the parameters assessed were not the same, but it is possible to infer some trend in sensitivity by looking at the differences between values obtained.

For seven out of eight compounds (abamectin, benomyl, carbofuran, copper, diazinon, dimethoate and lindane), Collembola showed a higher acute and chronic sensitivity than Isopodsa (several orders of magnitude higher). Indirect comparisons (looking also at chronic parameters) revealed a similar trend. The exception was L-cyhalothrin, where P. pruinosus was more sensitive than F. candida with effects on reproduction occurring at lower concentrations.

As indicated in Sections5 and 6.2.4, isopods are key drivers for several ecosystem processes, especially those linked to organic matter decomposition and nutrient cycling. Although they can take up chemicals by being in contact with contaminated soil (via the cuticle), the main route of exposure for most isopod species is in litter. Isopods can take up chemicals via contact with moist litter surfaces, but their exposure is higher when they feed on contaminated litter material (Peijnenburg et al., 2012).

When considering effects of PPPs on isopods exposed via soil, existing data show that their sensitivity is covered by other test species, e.g. the collembolan Folsomia candida (see above). However, the assessment of effects via exposure to contaminated litter, especially via food consumption, should be considered when designing a more appropriate hazard assessment of these compounds.

Despite the absence of a fully standardised ecotoxicity test (i.e. ISO or OECD guideline) on isopods, there is a vast experience in the literature on using them as model organisms for ecotoxicological evaluations of PPPs, ranging from assessing individual and population-level parameters (e.g. J€ansch et al., 2005; Morgado et al., 2016; Vink et al., 1995; Zidar et al., 2012) to ecotoxicogenomic studies (e.g. Costa et al., 2013a,b). Most published studies with PPPs focus on three species (Porcellio scaber, Porcellio dilatatus and Porcellionides pruinosus) and address lethal effects and effects linked to food consumption (measuring consumption, assimilation and assimilation efficiency) and their direct effects on energy allocation (measuring energy reserves) and, ultimately on growth (measuring biomass changes) at an individual level (Drobne et al., 2008; Ferreira et al., 2015; Ribeiro et al., 2001; Stanek et al., 2006; Zidar et al., 2012). Some studies also address behavioural parameters (Engenheiro et al., 2005; Loureiro et al., 2005, 2009; Santos et al., 2010). Although all these parameters can influence the onset of reproduction, number of offspring and, ultimately, population growth, it would be important to have more information on the direct effects of chemicals on reproductive parameters. In fact, the number of studies addressing direct effects on isopod reproduction is quite rare (e.g. J€ansch et al., 2005).

All these aspects, allied to the need to assess effects to key in-soil organisms having a relevant exposure to PPPs via consumption of litter debris, prompt the need to develop further a standardised test addressing both feeding and reproduction parameters. Despite the few papers on the optimisation of culture and test conditions (e.g. Caseiro et al., 2000), further research on the optimal isopod species, test design (e.g. test media, test duration, type of parameters), and litter material to use is needed. The extensive information already existing in literature could be a good starting point for a proposal for an ISO or OECD guideline needed to cover this group of key drivers via this particular and relevant exposure route to PPPs.

In conclusion, based on the available literature, F. candida may be protective for chronic effects on isopods when exposed via soil in the laboratory, taking into account an appropriate assessment factor.

However, Isopoda are key drivers for several ecosystem processes and the preferential route of exposure for most isopod species is litter on the soil surface. Therefore, the panel recommends development of a standardised test addressing both feeding and reproduction parameters, in order to assess effects to key in-soil organisms having a relevant exposure to PPPs via consumption of litter debris. The available information on isopods renders this a good candidate for developing an ISO or an OECD guideline.

Conclusion on the representativeness of Folsomia candida/Folsomia fimetaria and Hypoaspis aculeifer for arthropod soil invertebrates

The few available data indicate that F. candida and/or F.fimetaria could be representative of other springtails with regard to their toxicological sensitivity. Since hardly any data were found comparing the sensitivity of H. aculeifer with other soil mites, no conclusion can be drawn concerning its representativeness for other mite species. Even though with regard to the representativeness for the ecosystem, H. aculeifer seems important since it is the only predator in the test battery for which a standardised test protocol is available, an analysis of its toxicological sensitivity indicated that it showed a relatively low sensitivity compared with F. candida. Based on the available literature, F. candida may be protective for chronic effects on isopods when exposed via soil in the laboratory.

However, none of these studies considered an important exposure route for these animals, i.e.

contact in the litter on the soil. Therefore, no conclusion can be drawn on that respect. In order to take this important exposure route into account, the Panel recommends development of a standardised test addressing both feeding and reproduction parameters, to assess effects to key in-soil organisms having a relevant exposure to PPPs via consumption of litter debris.

9.2.2. Conclusion and recommendations for the choice of the standard

W dokumencie Scientific Opinion addressing the state of the science on risk assessment of plant protection products for in-soil organisms (Stron 116-122)