Effects of flow intermittency on Alpine streams functional processes: are
artificial flumes and field data comparable?
Gruppuso L.1,4*, Doretto A.2,4, Piano E.1,4, Falasco E.1,4, Bruno M. C.3, Bona F.1,4, Fenoglio S.1,4 1 DBIOS, University of Turin, Via Accademia Albertina 13, 10123 Turin, Italy
2 DISIT, University of Piemonte Orientale, Viale Teresa Michel 25, 15121 Alessandria, Italy 3 Department of Sustainable Agro-ecosystems and Bioresources, Research and Innovation Centre, Fondazione Edmund Mach, San Michele all’Adige (TN), Italy
4 Centro per lo Studio dei Fiumi Alpini (ALPSTREAM - Alpine Stream Research Center), Ostana (CN), Italy
*Corresponding author: Gruppuso Laura, laura.gruppuso@unito.it
Abstract
The disappearance of surface running water is becoming more and more frequent due to the effects of global climate change and growing human pressures. This phenomenon is also affecting several Alpine streams, whose regime is shifting from perennial to intermittent. However, the effects of flow intermittency on aquatic communities and ecosystem processes such as detritus decomposition have been poorly investigated, especially in field studies, due to the unpredictability of flow
intermittency and to the presence of other confounding environmental factors. To overcome these difficulties, mesocosm and artificial flumes experiments are often used in freshwater ecology, since they provide the possibility to control the effects of confounding variables that can occur in field research. Nevertheless, the results obtained from flumes simulations have to be weighed up and, if necessary, calibrated with field data. In order to evaluate the applicability of manipulative
experiments to natural conditions, we compared the results of a flumes simulation on CPOM processing with those obtained from field data. We compared an intermittent treatment and a control under permanent flow in: a) an artificial flume system; b) two Alpine rivers; and, in both experiments, we sampled the macroinvertebrate community and measured the leaf decomposition rates at approximately the same fixed time intervals. We obtained consistent results between the two experiments, and we observed that the percentage of remaining mass, the abundance of shredders, and the effect of the loss of water flow on macroinvertebrate community composition showed almost the same trend in both experiments. This study shows how flumes simulations can provide a valid proxy for naturally-occurring processes in stream ecosystems.
Keywords: leaf bags; droughts; climate change; leaf litter decomposition; benthic
macroinvertebrates 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35
Introduction
Because of global change, the occurrence of drought events in the near future is predicted to increase in both frequency and intensity (Lake 2000; Vicente-Serrano et al. 2014; Papadaki et al. 2016), and to increasingly affect streams that were previously perennial. In particular, the increase in water temperatures and the disruption of hydrologic cycles are expected to have remarkable effects on lotic habitats and on the composition of their communities, with consequences on metabolic instream processes (Ledger & Milner 2015; Abril et al. 2016; Falasco et al. 2018, 2020; Receveur et al. 2020). To date, the effects of droughts have been mainly investigated in the
Mediterranean ecoregion, where droughts are part of the natural hydrological cycle, and biological communities have evolved adaptations to survive the dry season and to rapidly recover when water flow returns (Bonada et al. 2007; Leigh et al. 2016). However, stream droughts are becoming an increasing problem in terms of economic loss, ecological damage and risks to human health (Bond et al. 2008) not only in Mediterranean streams, but also in larger areas of Europe, where Alpine streams are among the most threatened systems (Fenoglio et al. 2010; Huang et al. 2016). Because the increase of droughts in the Alpine area has been occurring in recent time, knowledge about the effects on stream and river biota is still in its early stages. In particular, some preliminary studies highlighted how Alpine macroinvertebrate communities probably lack strategies and adaptations to survive this hydrological stress. For example, EPT (Ephemeroptera, Plecoptera and Trichoptera) taxa decreased in intermittent sites, along with those taxa with long pre-imaginal life (large and semivoltine) which are not able to survive in dry conditions (Doretto et al. 2018; Piano et al. 2019a). Furthermore, non-flow events can also affect some specialized functional feeding groups, such as shredders, disturbing regular trophic interactions (Amundrud & Srivastava 2016) and causing the disruption of leaf litter breakdown processes (Ledger et al. 2011; Wenish et al. 2017). In addition, microbial organisms can be affected by water loss, making CPOM less palatable for macroinvertebrates (Boulton and Lake 1992). The effects of droughts are also mediated by the changes in flow characteristics: rheophilous taxa have been shown to decrease from perennial to intermittent sites, and lentic ones to increase, as the latter are able to overcome water fluctuation periods (Acuña et al. 2005; Pace et al. 2013; Chessmann 2015).
The increase of droughts in previously perennial streams will also likely alter most of the ecological processes, from primary instream production to allochthonous CPOM metabolism (Fenoglio et al. 2015; Pinna et al. 2016). Leaf litter decomposition plays a key role in nutrient cycling in streams (Petersen & Cummins 1974; Graҫa & Canhoto 2006): as this process includes leaching of soluble compounds, physical abrasion, microbial conditioning and invertebrate fragmentation activity, it depends on both intrinsic (i.e. litter quality) and extrinsic factors including temperatures and
dissolved nutrients (Tank et al. 2010; Chessman 2018). Intrinsic factors such as C:Nutrients content, tannin content and leaf texture can affect the decomposition process, slowing it down in case of poor quality and coriaceous leaf litter (Simon et al. 2018; Zhang et al. 2019). However, extrinsic factors can also play a key role in this process, especially during dry periods: variations in some of these parameters can affect bacterial and fungal activity, with indirect impacts on macroinvertebrate communities (Boyero et al. 2011; Follstad Shah et al. 2017). For example, leaf decomposition rate is usually faster in warm than in cold conditions, but it can differ between rivers at different altitudes or ecoregions. Decomposition rate has been proposed as an indicator of river ecosystem health and functionality; this approach relates to the food web theory, because disturbances make food webs less efficient in organic matter processing (Bonada et al. 2006). To our knowledge, the impacts of winter droughts on aquatic communities and ecosystem processes have been poorly assessed through field studies in Alpine and perialpine areas where allochthonous CPOM input in fall/winter is highest, decomposition occurs under very low temperatures, and flow reduction and 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60 61 62 63 64 65 66 67 68 69 70 71 72 73 74 75 76 77 78 79 80 81 82
winter droughts might cause leaf litter to be exposed to emersion-immersion cycles, affecting organic matter processing.
Although organic matter processing has been widely investigated in manipulative field studies, mainly by introducing standardized leaf bags in natural streams (Yoshimura et al. 2010), field studies can be difficult to perform because droughts are unpredictable, and studying their effects can be misinterpreted, due to the presence of a large number of confounding natural variables, such as the timing and frequency of non-flow periods, presence of refuges, and the co-occurrence of other anthropogenic disturbances (i.e. organic pollution, siltation, etc.). However, even if artificial streams experiments can be useful in eliminating all these confounding variables, they can represent a simplified situation that does not reflect what really occurs in natural conditions. Most of the studies conducted on leaf litter decomposition have been so far carried out either in the field (Corti et al. 2011; Calapez et al. 2014; Elias et al. 2015) or in artificial conditions such as microcosms and artificial flumes (Rumbos et al. 2010; Doretto et al. 2018; Santschi et al. 2018; Truchy et al. 2020). Therefore, to our knowledge, no studies have compared field and artificial flumes data on CPOM degradation. By collecting data obtained in a field study and a flumes simulation, our aim was to estimate if results of CPOM processing and macroinvertebrate community obtained in artificial flumes could simulate properly what really occurs in natural conditions. We were interested in evaluating the cumulative effects of flow intermittency compared to permanent flow and we hypothesized that: a) due to the correspondence of the leaf types used, independently from the type of experiment, leaf litter decomposition rate would be higher in the perennial sites; b)
macroinvertebrate communities would be affected by flow intermittency in both experimental conditions, reducing taxa richness (i.e. effect of the nestedness); c) the abundance of shredders would be linked to the presence of water flow; d) overall, even if in artificial flumes conditions are controlled, the results can be comparable to the ones obtained from the field studies.
Materials and methods
In the present study, two analogous experiments were carried out under different conditions. First, we performed a flumes simulation ina set of metal flumes (Fersina Stream, Eastern Italian Alps); second, we performed a field experiment in the Pellice and Varaita rivers (Piemonte, Western Italian Alps). The two experiments were comparable in duration (100 and 105 days respectively), quality of leaf litter used, beech (Fagus sylvatica) and oak (Quercus robur), respectively; in literature, both leaf types are defined as low quality ones (McKie & Malmqvist 2009; Sanpera-Calbet et al. 2009; Martinez et al. 2016; Simon et al. 2018; Santonja et al. 2019; Zhang et al. 2019) and the nature of the rivers examined (both Alpine streams). Each experiment is described in details in the following paragraphs.
Artificial flumes
The study was conducted in a set of open air metal flumes situated on the riparian zone of the right bank of a 2nd order stream (Fersina Stream, Trentino Province. 675836 E; 5104975 N).
The Fersina is a snowmelt-fed gravel bed stream approximately 14 km long, with a 171 km2 watershed area, and numerous small tributary streams flowing from lateral valleys. The
experimental setting consists of two 20 m long, 30 cm wide, 30 cm tall metal flumes. Each flume has a sluice gate to control discharge and connect them to a loading tank of 1 m3 volume, that is directly fed by water diverted from the stream. Sluice gates can be completely closed to block the incoming flow and simulate a drought. The flumes are filled to approximately the same depth with two layers of cobbles of ca. 10 cm diameter, and a layer of silt/sand/gravel which naturally collects 83 84 85 86 87 88 89 90 91 92 93 94 95 96 97 98 99 100 101 102 103 104 105 106 107 108 109 110 111 112 113 114 115 116 117 118 119 120 121 122 123 124 125 126 127 128
around the stones. The flumes were successfully used for several experiments targeting benthic macroinvertebrates and periphyton (e.g., Bruno et al. 2013, 2016; Grubisic et al. 2017, Doretto et al. 2018). The artificial streams were run at a baseflow of 5 l s-1, velocity 0.4 m s-1, from the beginning of April 2016, and left undisturbed at this same baseflow for 3 months previous to the experiment, in order to allow natural colonization by benthic invertebrates and periphyton by drift from
upstream sites and/or egg deposition. The experiment lasted 100 days: in flume C (i.e. Control, no drought), flow was maintained constant throughout the entire experimental period; in flume I (i.e., Intermittent treatment) we alternated 20 days of flow with 20 days of drought, repeated twice. In each flume we installed 50 leaf bags (weight 3.0 ± 0.1 g) made of coarse mesh (1 cm, in order to allow macroinvertebrate colonization), built with dry European beech (Fagus sylvatica) leaves, collected in September, air-dried for 15 days and stored in dark and dry conditions until the onset of the experiment. The 50 leaf bags were tied to two ropes, 25 leaf bags each, which were in turn fixed to the bottom of the flume. Leaf bags were deployed on 20 October 2016 and sampling was
conducted on five consecutive occasions, every 20 days (Table 1). On each sampling date, seven leaf bags were randomly removed in each flume. Leaf bags were inserted in a container, kept refrigerated and carried to the laboratory, where the leaf material from each bag was rinsed with distilled water to remove invertebrates and inorganic particles, and benthic invertebrates were sorted under a stereomicroscope and identified to the lowest possible taxonomic level following Campaioli et al. (1994, 1999) and Waringer & Graf (2011). Many early instars of insects were identified no further than to order or family level. The remaining leaf bag material was immediately air dried for 25 h and subsequently oven-dried (24 h, 80 °C) and weighted to the nearest 0.1 g, to determine dry mass and calculating leaf litter mass loss. Physical and chemical parameters (temperature, conductivity, pH, oxygen concentration) were measured before the simulations, in each experimental flume, by using a WTW© handheld Oxygen meter, pH meter, conductivity meter, and turbidity meter (WTW GmBH, Weilheim, Germany). These data are provided in Table 2.
Field experiment
The Pellice River, being approximately 53 km long and with a 975 km2 watershed area, is the largest left tributary of the Po River in the Cottian Alps area. The sampling station on this river (named “permanent” hereafter) was located near the Campiglione Fenile Village (366638E, 4964043N) and surface water flow was always present throughout the sampling campaign. The Varaita River is the first right tributary of the Po River, approximately 92.4 km long and with a 605 km2 watershed area. The sampling station along this river (named as “intermittent” hereafter) was located near Costigliole Saluzzo village (382886E; 4937757N) and flow intermittency events occurred during the sampling period. In each sampling station we deployed 60 leaf bags made of coarse mesh (1 cm, in order to allow macroinvertebrate colonization) of oak (Quercus robur) dry leaves (5.0 ± 1.0 g of leaves each, collected in October and stored in dark and dry conditions before being deployed). The sampling campaign began on 13 December 2018 and consisted of five
consecutive sampling dates (Table 1). Seven leaf bags were removed randomly every 21 days, stored in 80% ethanol and, in the laboratory, leaf material was separated from macroinvertebrates which were determined at family/genus level following Campaioli et al. (1994, 1999), Sansoni (1988) and Tachet et al. (1984, 2005), using a stereomicroscope. The leaves were rinsed with water to remove ethanol and then dried in an oven at 105° C for 24 hours; afterwards, dried material was weighed to determine dry mass and percentage of remaining leaf litter mass was calculated. 129 130 131 132 133 134 135 136 137 138 139 140 141 142 143 144 145 146 147 148 149 150 151 152 153 154 155 156 157 158 159 160 161 162 163 164 165 166 167 168 169 170 171 172 173 174
Physical and chemical parameters (temperature, conductivity, pH, oxygen concentration) were measured at each sampling date, with Hydrolab Quanta probe (Table 2). In addition, a Hobo temperature datalogger was deployed in each stream, to record temperatures every 6 hours throughout the experiment, and used as a proxy to assess the effective immersion period (Fig. 1).
Statistical analyses
Both datasets were analyzed with the same statistical analyses to better compare the results from the two experiments. CPOM mass loss (i.e., remaining mass, expressed as % of the initial weight) over time was visually examined by plotting the mean (±SE) percentage of remaining mass on each sampling date. Statistical differences in mass loss were tested only for the last sampling date: we chose to focus solely on the last sampling date because we were interested in evaluating the cumulative effects of flow intermittency compared to permanent flow, rather than short-term variation in the leaf-litter processing. A Mann-Whitney U-test was used to test differences in CPOM mass loss between the control and the intermittent flume and between the Pellice and Varaita rivers, for the artificial flumes and field experiments, respectively.
For each flume and/or river, differences among sampling dates (i.e., T0-T5) in the total taxa richness and abundance of shredders were statistically tested with a Kruskal-Wallis test and, when significant, pairwise comparisons were performed. Changes in the composition of macroinvertebrate communities associated with leaf bags in flumes or in the field were visually examined with a non-metric Multidimensional Scaling (NMDS) and were statistically tested for differences among “site” (i.e., permanent vs intermittent) and “sampling date” with a Permutational Analysis of Variance (Permanova). Macroinvertebrate abundances were log(x+1) transformed and the Bray-Curtis dissimilarity index was applied.
To better investigate the mechanisms responsible for the variation in community composition, we decomposed the total beta-diversity into its nestedness and turnover components using the approach suggested by Cardoso et al. (2020). Because we were interested in evaluating changes over time (i.e. along with the leaf-litter breakdown), macroinvertebrate taxa associated to leaf-bags on the first removal date were pooled together and considered as the reference communities for each flume and/or site. We calculated the Sorensen index of dissimilarity to compare total ꞵ-diversity, nestedness and turnover between the reference communities and each of the sampling dates (i.e. T1 vs T2, T1 vs T3 and so forth) for each flume/river.
All the statistical analyses (significance threshold = 0.05) were performed in R (R Core Team 2019), using basic functions and the following packages: vegan (Oksanen et al. 2015) for NMDS, and PERMANOVA, and BAT (Cardoso et al. 2020) for beta-diversity decomposition. Plots were drawn using the ggplot2 and ggpubr packages (Wickham 2016; Kassambara 2017).
Results
Drought events
In artificial flumes, the two simulated droughts lasted approximately 20 days, interspersed with 20 days of water flow each time. In the intermittent river, drought occurred at T2 (after 42 days), with most of the leaf bags completely emerged; emersion was confirmed by the datalogger, which 175 176 177 178 179 180 181 182 183 184 185 186 187 188 189 190 191 192 193 194 195 196 197 198 199 200 201 202 203 204 205 206 207 208 209 210 211 212 213 214 215 216 217 218 219 220
showed a sharp increase of temperature in those weeks as they were recording the air temperature (Fig. 1); water flow partially returned at T3 because of the presence of a small tributary stream and remained present but scarce until the end of the experiment. On the contrary, in the perennial river, water flowed for the entire sampling campaign. We therefore assumed that the Varaita River could be compared to the intermittent flume because of the similar water oscillations and length of dry events, while the Pellice River, where we did not observe flow intermittency events, could be compared to the control flume.
Leaf litter decomposition
We assessed the mass loss of the leaf bags during the whole experiment (number of days from T0, when leaf bags were placed in the riverbed) in both artificial flumes (Fig. 2a).
In the two artificial flumes mass loss had a similar temporal trend, even if it was more pronounced in the control flume. At the end of the experiment (100 days) the percentage of remaining mass was significantly lower (Mann-Whitney test: p = 0.008) in the control flume than in the intermittent one (Fig. 2b). There were on average eleven benthic taxa associated with leaf bags in the control flume at T1, and no statistical variation over time was observed in the succeeding sampling dates (Fig. 2c). Similarly, there were on average eleven taxa in the intermittent flume, but we recorded a progressive, albeit not significant, decline in taxa richness over time (Fig. 2d). The temporal patterns of shredders abundance also differed between the two flumes. In the control flume, the number of shredder specimens per leaf bag ranged from 0 to 25, without significant variation between sampling occasions (Fig. 2e). On the contrary, in the intermittent flume the highest abundance of shredders was recorded at T1; the number of macroinvertebrates belonging to this FFG significantly dropped in the following sampling dates and never approached the initial value (Fig. 2f).
In the field experiment, mass loss had almost the same temporal trend in the two rivers (Fig. 3a), but in the perennial site mass loss was more pronounced, similarly to what occurred in the flumes. In the permanent river at the end of the study (105 days) the percentage of remaining mass was lower (Mann-Whitney test: p = 0.002) than in the intermittent one (Fig. 3b). The two rivers had opposite trends in temporal variations of taxa richness: in the permanent one the average number of taxa significantly increased over time and at the end of the study (T5) taxa richness was more than four-fold higher than at T1 (Fig. 3c). By contrast, the opposite trend occurred at the intermittent river: the mean taxa richness was highest (five taxa) at T1, and then it significantly decreased over time, with a value of 3 taxa on the last sampling occasion (T5; Fig. 3d). Similar to what recorded for taxon richness, a significant increase in the number of shredders colonizing leaf bags over time (Kruskal-Wallis test = 21.224; df = 4; p < 0.001) was observed in the permanent river (Fig. 3e), while the shredders abundance in the intermittent one was lower and peaked at T3.
Macroinvertebrate community
Macroinvertebrate communities were investigated with a multivariate and a β-diversity analysis. In the artificial flumes, Permanova showed significant changes in the macroinvertebrate community composition in relation to the factors time, flume and their interaction (always p < 0.001). At T1 (after the first 20 days), the community of both flumes had approximately the same taxa
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composition; however, after 40 days (i.e., at T2), it began to change in the intermittent flume, while in the control flume the community was similar to the one recorded on the first sampling date (Fig. 4a). In the control flume, the community changes recorded at T2 were due to the effect of both nestedness (49%) and turnover (51%); the effect of turnover increased to 80% at T3 and 95% at T4 (Fig.4b). On the opposite, in the intermittent flume nestedness represented almost the 100% of contribution to beta diversity in all sampling occasions (Fig. 4c). β-diversity varied moderately over time in the control flume (increasing from 0.22 at T2 to 0.41 at T3), while it varied in a more remarkable way in the intermittent one (decreasing at T3 from 0.83 to 0.66, and increasing again at T4 to decrease at T5 from 0.98 to 0.60) (Fig. 4b, 4c).
In the two field sites we observed the same trend of the flumes: the community composition of both rivers was almost the same at T1 and began to change from T2 in the intermittent river, whilst it remained almost the same throughout the entire experiment in the permanent river (Fig. 5a). Differences were statistically significant over time and between rivers (Permanova: p < 0.001) and for the interaction between these two factors (Permanova: p< 0.004). In the permanent river community changes were due mainly to nestedness in all the sampling dates (95%, 70%, 90%, 100% from T2 to T5 respectively), with a little effect of turnover (Fig. 5b). Community changes were due mainly to the effect of nestedness in the intermittent river as well (85%, 65%, 80% and 95% from T2 to T5, respectively) (Fig. 5c). β-diversity, in the permanent river, decreased at T3 (from 0.17 to 0.09) and then increased at T4 to decrease again at T5 (0.53 and 0.14 respectively); in the intermittent river the trend was different and β-diversity decreased at T3 (from 0.83 to 0.47) and then increased at T4 and T5 (0.71 and 0.91 respectively). Considering the flumes and natural sites overall, total β-diversity varied moderately in the two control sites, while the temporal variations were more pronounced in the intermittent ones.
Discussion
The aim of this study was to compare the effects of flow intermittency on leaf litter breakdown and macroinvertebrates communities based on results obtained in an artificial flumes simulation and in a field experiment with similar setting, and to evaluate if the two types of experiments would provide comparable results. Indeed, CPOM decomposition was strongly affected by drought events with consistent patterns in both flumes and field experiments. Macroinvertebrates and degradation responses in the field slightly differed from those observed in the flumes because of the presence of natural variables which were controlled in the flumes (mainly, variable discharge in natural
conditions, constant discharge in flumes, and a reduced morphological variability in the flumes). However, a very similar pattern was detected, showing that hydrological intermittency and resulting surface water loss was the overriding factor determining biodiversity patterns and functional
responses, as already shown regarding temperate streams, for instance by Monroy et al. (2017).
Leaf litter decomposition
A large number of experiments have been carried out on leaf litter decomposition, mostly in naturally intermittent streams (Mora-Gómez et al. 2018; Fritz et al. 2019; Smeti et al. 2019). The results show that, in Mediterranean streams, summer drought events cause variance of litter processing rates (Monroy et al. 2016), and that the resilience to this perturbation is higher in high 265 266 267 268 269 270 271 272 273 274 275 276 277 278 279 280 281 282 283 284 285 286 287 288 289 290 291 292 293 294 295 296 297 298 299 300 301 302 303 304 305 306 307 308
order streams (Pinna & Basset 2004). Moreover, during intermittent phases, leaf decomposition is heterogeneous, because the shift from running waters to pools or dry stretches affects the microbial and fungal colonization of leaf litter (Abril et al. 2016). Temperature is also an important factor as in field experiments it strongly interacts with microbial breakdown, and consequently affects macroinvertebrate leaf litter decomposition (Mora-Gόmez et al. 2015; Correa-Araneda et al. 2020). In addition, the exposition of leaf litter to summer drought can change leaf litter chemical
composition, increasing nitrogen, lignin and cellulose content, with effects on the decomposition when flow returns (Mora-Gόmez et al. 2019).In the initial stage, leaf litter decomposition is led by the leaching phenomenon, with a loss of 20-25% of the total mass; after a few days the leaves become more palatable for shredders macroinvertebrates because of the conditioning action of bacterial and fungal activity (Fenoglio et al. 2019). Our results clearly suggest that intermittence slowed down leaf litter decomposition, because drought events and litter desiccation probably affected the conditioning process controlled by micro-consumers and clearly affected
macroinvertebrate community, especially the organisms which usually feed on CPOM (i.e.
shredders; Merritt & Cummins 1996). In fact, leaf bags showed the higher abundance of shredders in the perennial sites or in the sampling occasions where water flow was present; generally, abundance of shredders began to increase from T2, after the initial conditioning phase.
Regarding winter droughts, a small number of experiments have been carried out (Dai et al. 2006; Sun & Yang 2012), because this phenomenon is recent and difficult to predict. These few studies focus mainly on physico-chemical and climatic aspects, rather than community composition or decomposition processes; nevertheless, it has been shown that, because litter fragmentation by shredders is simulated by microbial activity, occasional freezing may constrain fungal diversity and their ecological functions (Fernandes et al. 2009) and delayed leaf decomposition accompanied by a decline in fungal biomass has been reported for the cold season, when temperatures are nearly 0°C (Nikolcheva and Bärlocher 2005). The results of our research, confirm that CPOM degradation is stronger and more efficient in perennial conditions even in winter.
Macroinvertebrate communities
At the beginning of both experiments, after 20-21 days of natural colonization, communities were almost the same in the control and intermittent site/flume; then, in the following sampling dates, the community composition in the intermittent site/flume was very different from the one present in the same sites before the drought, and the one of the perennial site/flume. Similar results have been reported in literature, showing that macroinvertebrates community composition is affected by dry periods (Giam et al. 2017; Doretto et al. 2020). For example, a study carried out using
hydrogeological indexes highlighted that community composition was influenced by drought (Kath et al. 2016). Intermittence strongly affected macroinvertebrate communities, probably because in Alpine streams, which historically were characterized by perennial flow, these organisms have not evolved an adaptation yet, and some of the taxa were not able to overcome the dry period (Lytle & Poff 2004). Biological communities of streams of mountain regions, such as in the European Alps, lack strategies and adaptation to survive such hydrological stress. In these environments, the occurrence of droughts is expected to impact the structure of lotic foodweb, with dramatic consequences on biodiversity conservation, river functionality and self-purification capacity of rivers. The occurrence of aseasonal droughts (i.e. in late fall/winter) can further impact the stream 309 310 311 312 313 314 315 316 317 318 319 320 321 322 323 324 325 326 327 328 329 330 331 332 333 334 335 336 337 338 339 340 341 342 343 344 345 346 347 348 349 350 351 352
ecosystems because low water levels can exacerbate the effects of low temperatures, increasing the amount and rate of freezing. It can be hypothesized that sensitive taxa were eliminated by
desiccation or moved into wet refugia during drought events, as reported in literature (Boulton 2003; Wood et al. 2010; Young et al. 2011; Bruno et al. 2020). In intermittent sampling sites we expected to find a lower percentage of EPT taxa, which are the most sensitive and dominant group in Alpine rivers (Dewson et al. 2007; Datry et al. 2014). Actually, our results show that community composition in those sites is dominated by Diptera, especially Chironomidae and, among
Ephemeroptera, one of the most abundant was Baetis sp., a plurivoltine organism of small size and fast larval development (tables regarding macroinvertebrates community composition in
supplementary materials). Some specialized functional feeding groups such as shredders and scrapers, are expected to decrease within intermittent traits, where the collector-gatherers increase instead (Acuña et al. 2015; Piano et al. 2019b). In our study, the abundance of shredders was higher in perennial sites where, in fact, leaf litter consumption was higher.
Nestedness and turnover analysis showed that the community which colonized the perennial sampling site, in both natural conditions and in the artificial flume, was richer and more stable than the one present in the intermittent site, where drought acted as a filter. The turnover process was recorded only 60 days after the beginning of the experiment, when some macroinvertebrates likely began to colonize CPOM after the conditioning phase, according to the ecological succession theory (Rossi 1985; Hieber & Gessner 2002). Moreover, the analysis highlighted how the nestedness pattern in the perennial sites was mainly due to the recruitment of new taxa, while in the
intermittent ones it was due to taxa loss. Furthermore, shredders abundance was higher where water flow was constant and drastically decreased in intermittent sampling sites; in field this FFG
increased in permanent river throughout sampling dates, while in the intermittent one it increased after the first drought, because water flow recovered due to the contribution of a right bank
tributary. Moreover, total β-diversity varied moderately in the two control sites in contrast with the wider temporal variation occurring in the intermittent ones indicates that macroinvertebrate
communities associated with the leaf bags were more dissimilar (i.e., less stable) in the two intermittent sites, as a consequence of the hydrologic variation.
Comparison of artificial flumes and natural conditions
In recent times, the use of experimental flumes of different typology has become a common tool to study the responses of biological communities to changes in hydrological, physical and chemical conditions (Bruno et al. 2013; Menczelesz et al. 2020 and references therein). In fact, the controlled experimental conditions of artificial flumes allow manipulating one or more factors of specific interest, thus disentangling relationships that are difficult to demonstrate in natural conditions. Indeed, Lamberti and Steinman (1993) suggested that a useful application of research conducted in artificial streams is the generation of testable hypotheses, which can then be validated in natural stream ecosystems. Nonetheless, in a recent review on the current knowledge on experimental streams, Menczelesz et al. (2020) highlighted some faults of these systems, such as size-thresholds below which the required level of biological complexity cannot be properly reproduced. In our study, the type and setting of artificial flumes appeared to perform well in assessing the effects of winter droughts on CPOM decomposition rates. In fact, CPOM degradation had the same trend in both the permanent river and the control flume, even if in controlled conditions the differences 353 354 355 356 357 358 359 360 361 362 363 364 365 366 367 368 369 370 371 372 373 374 375 376 377 378 379 380 381 382 383 384 385 386 387 388 389 390 391 392 393 394 395 396
between the two flumes (control and intermittent) were more pronounced than in the two rivers. Conversely, in the field, this pattern was more variable and more evident at the end of the experiment, possibly due to the higher hydrological and morphological variability. In any case, focusing on the last sampling date, the remaining mass in our leaf bags was lower in those sites where water was present during the whole experiment, regardless of the setting (artificial or natural).
Conclusions
To our knowledge, no study comparing field and mesocosm experiments in running waters has been carried out so far, especially in Alpine streams. In particular, the effects of drought events and the shifting from perennial to intermittent water flow in those streams, especially during winter, has been so far poorly investigated (Fenoglio et al. 2007; Bruno et al. 2020). Our comparison
highlighted that: a) leaf litter decomposition process is more efficient if water is present during the entire leaf litter processing time; b) macroinvertebrate communities that can be found in intermittent stretches are clearly different and less diverse than the ones present in perennial ones; c) shredder macroinvertebrates are more abundant in perennial reaches, where leaf litter is, as a consequence, used and degraded more efficiently. Although with few and expected discrepancies, our study shows that field and mesocosm data provide consistent results, because they give a similar pattern in their responses. This is extremely useful from the perspective of further investigating
intermittence and the increasing of drought events in Alpine streams, because the use of semi-artificial flumes can represent an effective and efficient way of simulating stress events or manipulating food webs.
Further work is needed, because intermittence in Alpine streams is a recent phenomenon that affects biodiversity and energetic pathways; moreover, learning how to effectively set and compare field work and artificial flumes is a strategic tool to better investigate droughts in a global climate change scenario.
Acknowledgements
Authors wish to thank F. Macchi, A. Maule, M. Rappocciolo, A. Tassone, L. Morchio, V. Sconfienza for their assistance in the field and laboratory activities. Alpstream (Alpine Stream Research Center) and Parco del Monviso are greatly acknowledged for their support during all the field activity. The Fersina flumes are managed by the Department of Civil, Environmental and Mechanical Engineering of the University of Trento together with Fondazione Edmund Mach. This work was realized within the framework of the PRIN NOACQUA “Risposte di comuNità e processi ecOsistemici in corsi d’ACQUA soggetti a intermittenza idrologica” project, code 201572HW8F, funded by Italian MIUR.
Authors contributions
Conceptualization: LG, AD; writing—original draft preparation: LG; formal analysis: LG, AD; laboratory work: LG, AD, MCB; all authors set the experimental designs, performed the field work, contributed critically to the drafts and gave final approval for publication.
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Legends to figures
Figure 1: Water temperature recorded in field sampling stations during the whole experiment.
759 760 761 762 763 764
Figure 2: Mass loss data in the artificial flumes: a) % of remaining mass ± SE during the whole
experiment; Black = Control, Grey = Intermittent. Boxplot showing: b) mass loss at T5 (after 100 days); c) taxa richness during on the different sampling dates in the control flume; d) taxa richness on the different sampling dates in the intermittent flume; e) shredders abundance on the different sampling dates in the control flume; f) shredders abundance during sampling dates on the different sampling dates in the intermittent flume. Black line: median value; lower and upper box edge: first and third quartile respectively; vertical lines: whiskers (± 1.5 interquartile distance). Letters above boxplot indicate significant differences based on pairwise comparisons.
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Figure 3: Mass loss data in the field; a) % of remaining mass ± SE during the whole experiment;
Black = Permanent river (Pellice), Grey = Intermittent river (Varaita). Boxplot showing: b) mass loss at T5 (after 105 days); c) taxa richness on the different sampling dates in the permanent river;
d) taxa richness on the different sampling dates in the intermittent river; e) shredders abundance on
the different sampling dates in the permanent river; f) shredders abundance on the different sampling dates in the intermittent river. Black line: median value; lower and upper box edge: first and third quartile respectively; vertical lines: whiskers (± 1.5 interquartile distance). Letters above boxplot indicate significant differences based on pairwise comparisons.
777 778 779 780 781 782 783 784 785 786 787
Figure 4: a) NMDS chart showing macroinvertebrate community composition in the artificial
flumes. Nestedness and Turnover for b) control and c) intermittent flumes. Numbers on the top indicate the total β-diversity: 0 is for a completely identical community and 1 for a completely different one. Community composition at T1 was used as a reference community to assess β-diversity and Nestedness and Turnover.
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Figure 5: a) NMDS chart showing macroinvertebrate community composition in the field sampling
sites. Nestedness and Turnover for b) permanent and c) intermittent rivers. Numbers on the top indicate the total β-diversity: 0 is for a completely identical community and 1 for a completely different one. Community composition at T1 was used as a reference community to assess β-diversity and Nestedness and Turnover.
797 798 799 800 801 802 803 804
Table 1: Sampling dates for both mesocosm and field experiments, and brief description of the
hydrological conditions in the treatment (i.e., intermittent) sites (one flume and one river, respectively for the mesocosm and field experiments).
Site Date Sample
Name
Event-flow conditions
Mesocosm 20-Oct-2016 T0 Leafpacks deployment. Flume I = flow
Mesocosm 10-Nov-2016 T1 Flume I = start first drought Mesocosm 30-Nov-2016 T2 Flume I = flow resumed Mesocosm 20-Dic-2016 T3 Flume I = start second drought Mesocosm 10-Jan-2017 T4 Flume I = flow resumed Mesocosm 30-Jan-2017 T5 Flume I = end of experiment
Field 13-Dec-2018 T0 Leafpacks deployment. Intermittent stream= flow
Field 03-Jan-2019 T1 Intermittent stream= flow
Field 24-Jan-2019 T2 Intermittent stream= drought Field 14-Feb-2019 T3 Intermittent stream= flow partially
resumed
Field 07-Mar-2019 T4 Intermittent stream= low flow Field 29-Mar-2019 T5 Intermittent stream= low flow, end
of experiment 805 806 807 808 809 810 811 812 813 814 815 816 817 818 819 820 821 822 823 824 825
Table 2: Physical and chemical parameters recorded during the sampling dates (T1 to T5) in both
experiments.
Site Parameter T0 T1 T2 T3 T4 T5
Flume C Water Temperature (°C) 11 5.1 3.1 3.8 1.5 1.9 Flume C Conductivity (µS cm-1) 148 86 79 85 82 83 Flume C Dissolved Oxygen (mg
L-1)
10.36 10.41 11.83 12.19 13.44 13.06
Flume C pH 7.96 8.13 8.09 8.04 7.98 8.03
Flume I Water Temperature (°C) 11 5 Dry 3.8 Dry 1.9 Flume I Conductivity (µS cm-1) 148.6 86.5 Dry 85.1 Dry 83.4 Flume I Dissolved Oxygen (mg
L-1)
10.25 10.38 Dry 13.01 Dry 13.40
Flume I pH 8.09 8.09 Dry 8.06 Dry 8.08
Pellice Water Temperature (°C) 3.65 2.34 4.25 3.70 7.88 8.89 Pellice Conductivity (µS cm-1) 140 145 160 154 160 164 Pellice Dissolved Oxygen (mg
L-1)
8.53 9.40 13.13 14.52 11.12 13.34
Pellice pH 7.78 8.16 8.03 7.62 7.86 7.93
Varaita Water Temperature (°C) 1.69 1.94 1.72 9.99 12.9 13.4 Varaita Conductivity (µS cm-1) 235 234 230 245 243 258 Varaita Dissolved Oxygen (mg
L-1) 12.54 12.60 12.32 12.29 12.92 12.96 Varaita pH 8.80 8.80 8.32 8.04 8.17 9.01 826 827 828 829
