4.2 Influence of different life cycles on species and OTU
detection
We hypothesized that community composition will differ depending on life
cycle differences between mero- and hololimnic taxa. Accordingly, we
found a strong effect of sampling season on all tested groups, except
Coleoptera. Species richness of merolimnic and hololimnic taxa were both
affected by sampling day but in a different way. In contrast to
merolimnic taxa, hololimnic taxa showed a more stable number of species
between autumn 2017 and spring 2018 likely due to the absence of
emergence events, as hypothesized. For the most species-rich hololimnic
group, Annelida, most species were detected in autumn with the most
species-rich taxa being assigned to the family Naididae. The high
diversity of Naididae probably comes from a high number of asexually
produced individuals, or through shedding of the sexually produced
individuals from autumn or dying of the adults after cocoon laying
(Learner et al., 1978). Contrary to our second hypothesis, we did detect
lower species numbers in summer also for the hololimnic group Annelida,
but the decrease was more steady instead of an abrupt drop in species
numbers after peaking as for merolimnic species.
In general, merolimnic species detection in the water depends on several
factors, for example time and duration of the flight period and
emergence, abundance of the species in the water, number of generations
per year, larval development and occurrence of dormancy/diapause.
Diptera were the most species-rich merolimnic order with most taxa
belonging to family Chironomidae. We detected a high diversity of
Diptera in spring and early summer which is consistent with the results
of other studies, implying a reduced diversity in winter and the highest
diversity in early summer before emergence, as emergence of chironomids
is correlated with high temperatures (Armitage et al., 2012; Bista et
al., 2017). The high number of species in spring is likely due to a high
growth rate preparing the larvae for emergence in summer, also
explaining the drop in detected species number for the later summer
month. For the species‑rich mero- and hololimnic groups, except
Annelida, no effect of sampling day on OTU richness was detected,
possibly due the high number of OTUs per group which can show distinct
responses to seasonal changes even within the same species. Contrary,
for Ephemeroptera, Plecoptera and Trichoptera, only OTU richness was
affected by sampling day, likely due to the lower numbers of species and
therefore smaller shifts in species richness over time. Plecoptera and
Trichoptera OTU richness showed one peak each, in winter and early
spring, which is likely linked to an increase in biomass that will
probably reach its peak right before emergence coinciding with our
second hypothesis expecting a low richness in summer. Plecoptera OTU
richness peaked a bit earlier than Trichoptera OTU richness which is
congruent with an on average earlier emergence of many Plecoptera
species compared to most Trichoptera species (Graf et al., 2008; Graf et
al., 2009; Graf et al., 2022a,b; Schmidt-Kloiber and Hering, 2015). By
comparison, Ephemeroptera OTU richness had two peaks, one in autumn and
one in late spring. Species details revealed that from 21 Ephemeroptera
species, ten have a bivoltine (mainly genus Baetis ), one a
flexible (Caenis beskidensis ) and two a semivoltine (genusEphemera ) lifecycle (Buffagni et al., 2009; Buffagni et al.,
2022; Schmidt-Kloiber and Hering, 2015). In contrast, the detected
Plecoptera and Trichoptera were mostly univoltine species.
Occurrence even between species within a merolimnic order differed
between seasons. The indicator species analysis revealed thatGlyphotaelius pellucidus is detectable in all winter and spring
months and absent in summer which coincides with the species having a
long flight period after emergence and a known dormancy (Graf et al.,
2008; Graf et al., 2022;
Schmidt-Kloiber,
A. and Hering D., 2015). In contrast Philopotamus ludificatushas also a long flight period and dormancy but was detected less
frequently but had similar read numbers as G. pellucidus (Graf et
al., 2008; Graf et al., 2022; Schmidt-Kloiber and Hering, 2015). As
neither differences between life cycle characteristics, nor read numbers
were present, the less frequent detection of P. ludificatus was
likely because the species was rarer at the sampling location or because
of other unknown factors influencing the detectability of the species.
Other species like the Ephemeroptera Baetis rhodani andBaetis vernus are detectable throughout the year with B.
vernus being less frequently detectable than B. rhodani . The
more frequent detection of B. rhodani can be explained due to the
species being also sometimes trivoltine and B. vernus being only
known for a bivoltine life cycle (Buffagni et al., 2009; Buffagni et
al., 2022; Schmidt-Kloiber and Hering, 2015) and therefore, larvae ofB. rhodani are probably more frequent. Additionally the OTUs
detected belonging to species B. vernus differed in their
seasonal occurrence with one OTU only occurring in autumn. It is known
that different OTUs can show distinct responses to environmental changes
(Beermann et al., 2018), and for the B. vernus group cryptic
diversity has been recorded (Ståhls & Savolainen, 2008). This
strengthens the assumption that the differences in detection between the
OTUs is based on different responses to environmental changes and
therefore an underestimation of the diversity within B. vernus . A
limitation in using eDNA to assess patterns of seasonality is the
persistence of eDNA in the environment for up to several days or weeks,
and apart from that, the uncertainty of capturing living or dead cells
(Dejean et al., 2011; Pilliod et al., 2014; Thomsen et al., 2012a,b).
Nonetheless, our results demonstrate that the patterns we found are
consistent with the phenology of the different taxa thus further
encouraging that the DNA we detected mostly originated from living
organisms.