organic material taken directly from the river as a natural source of fungi and bacteria
(e.g. Zubrod et al. 2015, 2017) (see Table 2). Leaves can also be directly conditioned
in situ by placing them in small nets/bags that are suspended in a river and retrieved
after a specific number of days (Alonso et al. 2009; De Castro-Català et al. 2017;
Forrow and Maltby 2000; Graça et al. 2001; Zubrod et al. 2015) (see Table 2).
Although river water might reproduce natural environmental conditions, it is
sometimes contaminated, and this might have an impact. When river water is
used, a chemical breakdown of the water should be undertaken and reported along
with the study findings, so that any contamination is transparent. It is especially
important to disclose if the river water is contaminated with the substance(s) under
investigation in the study. If the test substances are present in the river water, the
organism could be exposed to that concentration and also the experimental dose.
Therefore, the organisms’ response would not be a true reflection of the test
concentration(s) but instead the reported dose combined with the concentration
found in the river water. For example, contaminants might be absorbed onto the
leaf surface and passed onto the organisms, or they could be released into the media,
which might happen during the acclimation period and/or during the experiment
itself, resulting in a compromised feeding activity. Therefore, the observed findings
might be an indirect effect, due to the leaf quality and not as a direct result of the
contaminant being tested.
The conditioning process usually takes around 2 weeks, but there are clear
differences in the literature about this stage (see Table 2). The process ranges from
a few days (Alonso et al. 2009), to several weeks (Blarer and Burkhardt-Holm 2016),
and up to months (Danger et al. 2012) (see Table 2). When Gammarus spp. are
offered a choice between leaves that have been conditioned for different periods of
time, they prefer those that have been conditioned for the longest (Agatz and Brown
2014; Bird and Kaushik 1985). Consequently, experiments (in situ or ex situ) that
provide the organisms with leaves that have been conditioned for a short or longer
time period could potentially underestimate or overestimate the actual feeding
activity of Gammarus spp.
It has been demonstrated that conditioned leaf material is more palatable (Agatz
and Brown 2014; Graça et al. 1993b) and that different species of leaves (i.e. Acer
spp. and Ulmus spp.), depending on the conditioning stage, might be more or less
palatable compared to the others (Bird and Kaushik 1985). Consequently, it could be
argued that it is impossible to compare experiments where organisms have been fed
with organic material that has been conditioned for different periods of time.
Organisms fed on leaves that have been conditioned for 1 week will probably eat
less than those fed with the same leaves conditioned for 3 weeks, and leaf unpalatability might be mistakenly attributed to contaminant exposure. It has also been
identified that Gammarus pulex fed with unconditioned leaves have a considerably
lower respiration rate (Graça et al. 1993b).
Depending on the methodology used, the conditioning process might take place
in different phases. There are studies where the leaves are provided to the organisms
directly after the conditioning process (Bärlocher and Kendrick 1973a; Bloor 2010;
Bundschuh et al. 2009; Forrow and Maltby 2000; Newton et al. 2018), whereas in
Feeding Behavioural Studies with Freshwater Gammarus spp.: The. . .
23
(e.g. Zubrod et al. 2015, 2017) (see Table 2). Leaves can also be directly conditioned
in situ by placing them in small nets/bags that are suspended in a river and retrieved
after a specific number of days (Alonso et al. 2009; De Castro-Català et al. 2017;
Forrow and Maltby 2000; Graça et al. 2001; Zubrod et al. 2015) (see Table 2).
Although river water might reproduce natural environmental conditions, it is
sometimes contaminated, and this might have an impact. When river water is
used, a chemical breakdown of the water should be undertaken and reported along
with the study findings, so that any contamination is transparent. It is especially
important to disclose if the river water is contaminated with the substance(s) under
investigation in the study. If the test substances are present in the river water, the
organism could be exposed to that concentration and also the experimental dose.
Therefore, the organisms’ response would not be a true reflection of the test
concentration(s) but instead the reported dose combined with the concentration
found in the river water. For example, contaminants might be absorbed onto the
leaf surface and passed onto the organisms, or they could be released into the media,
which might happen during the acclimation period and/or during the experiment
itself, resulting in a compromised feeding activity. Therefore, the observed findings
might be an indirect effect, due to the leaf quality and not as a direct result of the
contaminant being tested.
The conditioning process usually takes around 2 weeks, but there are clear
differences in the literature about this stage (see Table 2). The process ranges from
a few days (Alonso et al. 2009), to several weeks (Blarer and Burkhardt-Holm 2016),
and up to months (Danger et al. 2012) (see Table 2). When Gammarus spp. are
offered a choice between leaves that have been conditioned for different periods of
time, they prefer those that have been conditioned for the longest (Agatz and Brown
2014; Bird and Kaushik 1985). Consequently, experiments (in situ or ex situ) that
provide the organisms with leaves that have been conditioned for a short or longer
time period could potentially underestimate or overestimate the actual feeding
activity of Gammarus spp.
It has been demonstrated that conditioned leaf material is more palatable (Agatz
and Brown 2014; Graça et al. 1993b) and that different species of leaves (i.e. Acer
spp. and Ulmus spp.), depending on the conditioning stage, might be more or less
palatable compared to the others (Bird and Kaushik 1985). Consequently, it could be
argued that it is impossible to compare experiments where organisms have been fed
with organic material that has been conditioned for different periods of time.
Organisms fed on leaves that have been conditioned for 1 week will probably eat
less than those fed with the same leaves conditioned for 3 weeks, and leaf unpalatability might be mistakenly attributed to contaminant exposure. It has also been
identified that Gammarus pulex fed with unconditioned leaves have a considerably
lower respiration rate (Graça et al. 1993b).
Depending on the methodology used, the conditioning process might take place
in different phases. There are studies where the leaves are provided to the organisms
directly after the conditioning process (Bärlocher and Kendrick 1973a; Bloor 2010;
Bundschuh et al. 2009; Forrow and Maltby 2000; Newton et al. 2018), whereas in
Feeding Behavioural Studies with Freshwater Gammarus spp.: The. . .
23