Comparison of Phenotypic Traits and Fitness for Three Populations of Austropotamobius Torrentium (Schrank, 1803)

BIHAREAN BIOLOGIST 11 (2): 71-78 ©Biharean Biologist, Oradea, Romania, 2017 Article No.: e171204 http://biozoojournals.ro/bihbiol/index.html

Comparison of phenotypic traits and fitness for three populations of Austropotamobius torrentium (Schrank, 1803)

Marius-Ioan GROZA* and Vioara MIREȘAN

Department of Fundamental Sciences and Biotechnologies, Faculty of Zootechnics and Biotechnologies, University of Agricultural Sciences and Veterinary Medicine, 400372 Cluj-Napoca, Romania. *Corresponding author, M.I. Groza, e-mail: [email protected]

Received: 10. February 2017 / Accepted: 14. June 2017 / Available online: 17. June 2017 / Printed: December 2017

Abstract. Changes in environment induce adaptive variation in phenotypic traits of individuals. Therefore, isolated populations can be accurately defined by specific divergent characteristics. Using morphometric measurements and individual weight we assessed body fitness and specific features for three geographically isolated populations of Austropotamobius torrentium in Romania. All populations had a good ecological condition with specific differences between sexes. Also, small differences occur between the condition of the populations probably, due to specific habitat features. We found significant differences between chelae and cephalothorax size in relation to total length of individuals for all three populations. These differences were seemingly correlated with altitude, water velocity and shelter stability. In-habitat distribution of individuals is linked to their size. Larger individuals occupy the most stable shelters all across the studied sections. Also, higher water flows were populated mostly by the largest individuals, probably due to their adaptability to higher currents. The present study contributes to the knowledge on ecological fitness, morphometric variation and in-situ scattering of A. torrentium. This is an important step in evaluation of conservation measures needed for the protection of the previous mentioned species in Romania.

Key words: crayfish, cephalothorax, shelter stability, water velocity, altitude.

Introduction spreading from the underground pools that sheltered individuals from the cold during the Pleistocene (Pârvulescu et Long term isolation between populations leads to divergent al. 2013) and being later limited by the interaction with traits in individuals that occur as optimum favorable adapta- Astacus astacus and water quality (Pârvulescu & Zaharia tion to changes in environment (Dugas et al. 2015). Thus, 2013, Pârvulescu et al. 2013, Pârvulescu & Zaharia 2014). phenotypic traits in animals tend to vary with differences in Pârvulescu et al. (2013) also found the species to be almost habitat features (e.g., Li & Denny 2004, Bower & Piller 2015, completely isolated by the other 3 species of crayfish present Lighten et al. 2016, Schalk 2016), occurring in some cases here (Astacus astacus, Astacus leptodactylus and Orconectes even before genome modifications (Imre et al. 2002, Aubret limosus) and divided into two geographically distinct meta- et al. 2004, Scoville & Pfrender 2010). Thereby, earlier varia- populations. tion in shape and morphometry can occur due to stress re- Various studies on morphometry of A. torrentium were lated pressure (West-Eberhard 2003, Badyaev 2005). Various published in the past (e.g. Streissl & Hödl 2002a, Maguire & studies observed changes determined by shifts in environ- Klobucar 2011, Vlach & Valdmanova 2015), but information mental conditions (Swain & Foote 1999, Lighten et al. 2016) dealing with the Romanian populations are scarce. Thus, our or the presence of predators (van Buskirk &Schmidt 2000, goal was to assess and compare the ecological conditions of Auld & Relyea 2011, Schalk 2016). Consistent to this phe- 3 different populations of A. torrentium and to evaluate mor- nomenon, crayfish also express phenotypic plasticity related phometric changes and in-situ distribution of individuals to its environment (e.g., Griffith et al. 1996, Sint et al. 2005, driven by habitat features. We expected to find important Sint et al. 2007, Haddaway et al. 2012, Perry et al. 2013, differences in morphometry and fitness of individuals as a Helms et al. 2015, Dakić & Maguire 2016). result of their long term geographical isolation. Additionally, Occurrence of stone crayfish (Austropotamobius torren- we also assumed a positive influence of habitat characteris- tium) in Europe is strongly related to high water quality tics in phenotypic traits expression. (e.g.,Vlach et al. 2009, Pârvulescu et al. 2011, Svobodová et al. 2012, Pârvulescu & Zaharia 2013) and low anthropogenic influence (Füreder et al. 2006). Small distance movement of Materials and methods individuals due to a more sedentary behavior (Kadlecová et Habitats al. 2012) narrows its range further, making the species more We chose three different streams (Fig. 1) for our study: Valea Satului, susceptible to habitat alteration and risk from invasive spe- Aninoasa and Valea Aușoarelor located in three different hydro- cies (Dyer et al. 2013). Furthermore, added influence of habi- graphical basins (Tab. 1). Minimum distance between locations is tat features and predation seem to affect in situ distribution more than 120 km. For each stream we chose a 1 km section, 500 met- and density within populations (Vlach et al. 2012). All these ters away from the nearest village. We chose upstream segments of factors lead to a patchy distribution of the species (Kouba et the streams which were not affected by human industrial activities. al. 2014) imposed by physical barriers, human activity and In every segment we inspected for shelters and examined the interior of each shelter by hand. When a crayfish was found, we noted GPS habitat degradation (Pârvulescu et al. 2016). An uneven dis- coordinates, altitude, substrate type in the surrounding area of the persal is also observed in Romania, where A.torrentium oc- shelter, water velocity near the entrance of the shelter and the maxi- curs only in the karstic regions of the Western Carpathians mum water velocity across the stream, along the same line with the (Pârvulescu 2009, Pârvulescu et al. 2013, Pârvulescu & Za- shelter. GPS coordinates and altitude were measured using an Etrex haria 2013). This scattering seems to be the result of former 30 unit from Garmin (Garmin, Garmin Ltd., Lenexa, Kansas, United

72 M.I. Groza et al.

Table 1. Habitat characteristics (Alt: altitude; Subs: substrate; SGP - sand, gravel, pebbles; water velocities: mean water velocity, maximum water velocity and gallery mean water velocity; LS-low stability, MS-medium stability, HS-high stability).

Water velocities % of observed occupied shelters Stream Alt. (m) Subs Mean (SE) Max G Mean (SE) LS MS HS Aninoasa 732-790 SGP 35.4(2.2) 61.1 16.8(2.9) 14.29 47.62 38.09 Valea Aușoarelor 350-486 SGP 28.4(2.9) 61.1 7.6(2.5) 16.67 20 63.33 Valea Satului 142-334 SGP 12.6(1.8) 30.0 3.1(0.9) 11.54 53.85 34.61

healthy, individuals with missing or regenerated chelae were not considered. Only data regarding the left chela was examined due to high similarity between both chelae in crayfish (Maguire & Dakic 2011). We used log-linear regression to calculate the correlation between total length and weight for males, females and both sexes for each population. We used the exponent’s (b) value to evaluate growth isometry and students t-test (ts=(b-3)/Sb; b-slope, Sb- standard error of b) function to evaluate the significance of the result (Datta et al. 2013). For a better evaluation and comparison between populations condition, we also calculated Fulton’s Condition Factor (FCF). For this we used the formula: FCF=Wwx100/L3, where Ww is the wet weight of the animal in grams and L is the total length of the animal in cm (Streissl & Hödl 2002a). For comparison between FCF’s index of populations and sexes we used One-way ANOVA test. Spearman was used to test correlation between morphometric features, sex of the crayfish and different characteristics of the habitats.

To find differences between males and females within the same Figure 1. Map with the GPS coordinates from the middle of the population we used Mann-Whitney U test. Kruskal-Wallis test was stream segment used to compare the features among habitats.

States). Substrate type was visually approximated in order to create a Results substrate coarseness ranking score, according to Gibson et al. (1998). We recorded the presence of a variety of boulders, large areas with a combination of sand, gravel and pebble and small patches with silt Population’s fitness and clay. We only considered sand, gravel and pebble as substrate There are differences in allometry of individuals. Length- for our analysis as they constitute more than 95% of the stream bot- weight relationship equation showed a positive allometric tom in the shelter proximity (an area of 1 squared meter). For meas- growth (b>3) in all males and isometric growth (b=3) in all urements of the water velocity we used a JDC Flowatch flowmeter females. Overall, we observed an isometric growth in Ani- (JDC Electronic SA, Waadt, Switzerland). Using these measurements, noasa and Valea Aușoarelor populations and an allometric we calculated the mean of the maximum speed values in the river. growth for the population from Valea Satului. There is a We also determined the ratio between water velocity near the entrance of the shelter and the maximum water velocity across the strong correlation between weight and length of all the indi- stream for each individual for a better understanding of crayfish viduals (Spearman, r>0.981, p<0.001). More than 90% of the preference towards water speeds near their refuge. Shelter stability variation in weight is explained by the variation in length for was also recorded in 3 different categories: low stability (LS), me- all specimens, independent of location. Fulton’s condition dium stability (MS) and high stability (HS) (according to Pârvulescu factor (FCF) differs significantly between sexes in all locali- et al. 2016). Thus, macrophytes, sand and gravel were considered as ties (ANOVA, all p<0.021), with the larger index belonging LS, smaller unstable rocks, tree roots and shallow galleries were con- to the males (Tab. 3). sidered as MS and stable rocks and deep galleries were treated as HS shelters. However, there is only a small positive difference in FCF between males and females from Valea Aușoarelor (9.38%). Animals In contrast, differences in FCF between sexes from the other In this study, a total of 106 individuals of A. torrentium (Tab. 2) were two localities are more than 21% in favor of males. Differ- recorded. We measured each individual using a digital caliper (0.1 ences in FCF are also significant between populations mm accuracy) for total length (TL) from tip of the rostrum to the dis- (ANOVA, F=4.289, p=0.016). Post hoc test (LSD) show no tal margin of the telson, cephalothorax length (CL) from post-orbital significant difference in Fulton’s condition factor (FCF) be- edge to the middle of cephalothorax-abdomen junction, and chelae tween Valea Aușoarelor and Aninoasa populations (p=0.862) length (ChL) from the tip to the base of the claw. To better under- stand differences between populations we divided the length of but significant difference between these two populations and cephalothorax and chelae with the total body length (CL/TL and the population from Valea Satului (all p<0.018) (Tab. 3). ChL/TL, respectively) and further used these indices in our analysis. We found specific differences between sexes. Overall, We also recorded wet weight (Ww) for each individual using a digi- there is a significant difference between males and females tal coin scale (0.01 g accuracy). We took all the samples in a single among cephalothorax length, chelae length, weight, CL/TL month (September 2015) as length and weight of the crayfish may ratio and ChL/TL ratio (Mann-Whitney U, all p<0.047). vary depending on the season (Lindqvist & Lathi 1983). Males have generally: longer cephalothorax (Spearman,

Statistics r=0.199, p=0.047), longer chelae (Spearman, r=373, p=0.0001), larger weight (Spearman, r=0.233, p=0.02) and higher CL/TL

For the statistical analysis, we chose only individuals that were

Comparison of phenotypic traits and fitness for three populations of Austropotamobius torrentium (Schrank, 1803) 73

Table 2. Total number of males and females and average measurements (TL – total length, CL – cephalothorax length, ChL – chelae length, Ww – weight; all values were expressed as mean (SE)).

Location Sex Nr. of ind. TL (mm) CL (mm) ChL (mm) Ww (g) CL/TL (%) ChL/TL (%) Aninoasa M 22 68.99(2.24) 31.12(1.36) 30.42(1.74) 16.64(2.03) 44.95(0.95) 43.49(1.14) F 20 70.06(2.37) 29.55(0.96) 24.39(0.95) 13.01(1.12) 42.42(0.87) 34.70(0.41) Valea Aușoarelor M 14 62.79(5.57) 26.42(2.51) 26.26(3.42) 14.70(3.24) 41.71(0.42) 39.82(1.92) F 17 62.00(3.86) 24.76(1.42) 21.17(1.57) 10.67(1.40) 40.40(0.68) 33.65(0.64) Valea Satului M 13 72.47(3.36) 29.01(1.53) 32.45(2.53) 17.84(2.66) 39.89(0.29) 43.98(1.53) F 14 66.06(3.93) 24.65(1.61) 23.34(1.81) 10.57(1.56) 37.12(0.39) 34.74(0.82)

Table 3. Length-weight relationship amongst sexes and populations (r2:coefficient of determination; b: slope; Fulton’s condition factor FCF; I: isometric growth; A(+): positive/negative allometric growth)

Location LogW= Loga + bLogL r2 b FCF Growth Aninoasa M LogW= -5.353+3.551LogL 0.973 3.551 4.567 A(+) F LogW= -4,123+2.825LogL 0.962 2.825 3.605 I All LogW= -4.650+3.141LogL 0.905 3.141 4.109 I Valea M LogW= -4.821+3.255LogL 0.994 3.255 4.305 A(+) Aușoarelor F LogW= -4.162+2.860LogL 0.991 2.86 3.901 I All LogW= -4.499+3.050LogL 0.987 3.05 4.083 I Valea Satului M LogW= -4.967+3.320LogL 0.987 3.32 4.239 A(+) F LogW= -4.684+3.103LogL 0.995 3.103 3.190 I All LogW= -5.014+3.315LogL 0.963 3.315 3.695 A(+)

Figure 2. Regression lines of weight, cephalothorax length, chelae length vs total length (■ – males; ○ – females).

(Spearman, r=0.385, p<0.0001) and ChL/TL ratios (Spear- ferences were shown only between Valea Aușoarelor and the man, r=0.698, p<0.0001), respectively. Within populations, other two populations in ChL/TL ratio (Mann-Whitney U, sex-related morphometric differences are slightly dissimilar all p<0.046). Populations of Aninoasa and Valea Satului (Tab. 4, Fig. 2). were very similar in this regard (Mann-Whitney U, U=559, We also found significant differences between popula- p=0.922). tions in CL/TL and ChL/TL ratios (Kruskal-Wallis, both p<0.033) (Tab. 2, Fig. 3). Pairwise comparison showed sig- Habitat characteristics nificant differences between all populations in CL/TL ratio The studied streams had a very similar substrate with a ran- (Mann-Whitney U, all p<0.006). Concomitant, significant dif- domly combination of sand, gravel and pebbles (Kruskal- 74 M.I. Groza et al.

Table 4. Diferences between males and females within populations. Males are positively correlated with higher values of morphometric measurements. Significant correlations (p<0.05) are marked with bold. (CL – cephalothorax length, ChL – chelae length, Ww – weight).

Morphometric Valea Valea Aninoasa Measurements Satului Ausoarelor CL r=0.94 r=0.338 r=0.145 ChL r=0.382 r=0.495 r=0.192

Ww r=0.157 r=0.381 r=0.159 CL/TL r=0.342 r=0.799 r=0.413 ChL/TL r=0.865 r=0.723 r=0.471

Figure 4. The correlation between the total length (TL), cephalothorax length (CL) of crayfish, water velocities at the entrance in the gallery (ENT), maximum water velocity nearby (MAX) and altitude of habitats.

Figure 5. Ratio of shelters within habitats (LS – low stability, MS – medium stability, HS – high stability)

Figure 3. Body ratios comparison between populations. Lines in lui and the other 2 streams (Kruskal-Wallis, all p<0.001). The boxes represent medians, bottom and top ends of the boxes rep- difference in mean water velocity between Valea Aușoarelor resent the first and third quartile, respectively, whiskers repre- and Aninoasa is not statistically significant (Kruskal-Wallis, sent 1.5 times interquartile range, dots represent outliers. χ2=12.28, p=0.074). Overall, average values of water velocity

at the gallery entrances do not exceed 37.6% (±4.08SE) of av- Wallis, χ2=3.888, p=0.143). However, altitude and mean wa- erage maximum speed of the flow nearby. The ratios be- ter velocity are significantly different between streams tween these velocities are significantly different between lo- (Kruskal-Wallis, all p<0.024) (Tab.1). Pairwise analysis also cations (Kruskal-Wallis, χ2=6.499, p=0.039), with Aninoasa showed significant differences between all streams (Kruskal- having the larger ratio of 0.47 (±0.07SE). Wallis, all p<0.011). In addition, there is a strong correlation We also analyzed the number of low, medium and high between altitude, stream mean water velocity, the velocity of stability shelters for each location (Tab. 1, Fig. 5). Although the water in front of the occupied shelters and maximum the number of shelters differs between streams, we didn’t water flow nearby (Spearman, all p<0.004). Thus, some dif- find statistically significant differences between locations ferences regarding water speeds near shelters occur between (Kruskal-Wallis, all p>0.052). locations (Fig. 4). Water velocity at the entrance of the shelters is significantly different in Aninoasa compared to Valea The crayfish morphometry vs habitat features Aușoarelor and Valea Satului (Kruskal-Wallis, p<0.037). This Overall presence of males and females in the streams is not feature had a higher mean for Aninoasa (Tab. 1). Difference correlated with altitude, water velocities or stability of shel- between Valea Aușoarelor and Valea Satului is not statisti- ter (Spearman, all p>0.108). However, females from Valea cally significant (Kruskal-Wallis, χ2=3.21, p=0.661). Maxi- Aușoarelor and Valea Satului can be found in shelters closest mum water velocity differ significantly between Valea Satu- to higher water flows (Spearman, r=0.410. p=0.022 and

Comparison of phenotypic traits and fitness for three populations of Austropotamobius torrentium (Schrank, 1803) 75

Figure 6. Effect of size and weight on choosing different type of shelter (low, medium or high stability). Lines in boxes represent medians, bottom and top ends of the boxes represent the first and third quartile, respectively, whiskers represent 1.5 times interquartile range, dots represent outliers.

r=0.395, p=0.041, respectively). We found significant correla- (r=0.438, p=0.022 and r=0.518, p=0.006). tions between morphometrics of individuals and habitat features. Thus, total length and weight are positively correlated with shelter stability (Spearman. r=0.364. p=0.0002 and Discussion r=0.409, p<0.0001) and maximum water velocity near their shelter (Spearman, r=0.204, p=0.042 and r=0.210, p=0.036). A Population’s condition significant positive correlation was also found between che- All three populations of A. torrentium had a good fitness. lae length and shelter stability (Spearman, r=0.409, p<0.0001) However, we found differences between populations in in- (Fig. 6). Moreover, cephalothorax length is positively corre- dividual conditions and morphology. Aninoasa and Valea lated with altitude (Spearman, r=0.250, p=0.012), maximum Aușoarelor individuals had a very close FCF. The situation is water velocity near the shelter and speed flow at the en- similar even when comparing males and females between trance of the shelter (Spearman. r=0.224, p=0.025 and these populations. A significantly smaller FCF was obtained r=0.248, p=0.013) (Fig. 4) as well as shelter stability (Spear- for Valea Satului population. Crayfish fitness usually fluctu- man, r=0.360, p=0.0002) (Fig. 6). No correlation was found ates, depending on a variety of environmental factors. Steissl between crayfish morphometrics and gallery entrance water & Hodl (2002a) and Maguire & Klobucar (2011) found sig- velocity / maximum water velocity nearby ratio (Spearman, nificantly increased fitness in populations inhabiting sec- all p>0.053). Overall, all analyzed traits are positively corre- tions of streams with high flow heterogeneity and substrate lated with shelter stability (Spearman, all p<0.048). Thus, diversity and increased availability of potential shelters. In larger crayfish seem to occupy the most stable refuges. contrast, anthropogenic intervention casts a negative influ- Moreover, crayfish most commonly used medium and high ence in the condition of a population (Fevolden & Hessen stability shelters. Only an average of 14.17% (±1.48SE) of all 1989, Streissl & Hödl 2002a, Maguire & Klobucar 2011, individuals used low stability shelters (Tab. 1). Etchinson et al. 2012). Even though anthropogenic influence In Aninoasa, we didn’t find correlations between TL, CL, is less susceptible for these populations, limited growth

ChL, Ww and altitude, gallery and maximum water velocity could also be influenced by environmental conditions like (Spearman, all p>0.054). Instead, we found significant posi- low temperatures, high water flows, short growing season tive correlations between CL/TL ratio and altitude and duration, limited food availability, high competition, spread maximum water velocity near the shelter (Spearman, all of parasites or genetics of the population (Flint 1975, Olsson p<0.034). We didn’t find significant correlation between et al. 2008, Perry et al. 2013, Endrizzi et al. 2013). Since these CL/TL ratio and shelter stability (Spearman, r=0.095, factors weren’t the focus of our study, a more comprehen- p=0.548). sive long term study on larger populations is needed for In Valea Aușoarelor the morphometric measurements conclusive correlations in this regard. FCF also differ be- are not correlated with altitudes, or water velocities (Spear- tween sexes with males having better conditions, which is man, all p>0.081). Shelter stability is not correlated with consistent with other studies on stone crayfish condition CL/TL (r=-0.031, p=0,867), but correlated with ChL/TL (e.g. Streissl & Hödl 2002a, Maguire & Klobucar 2011). (r=0.420, p=0.019). Tracking changes in weight and length of individuals re- In Valea Satului the morphometrics are not correlated vealed the ratio of growth amongst different sized crayfish. with altitudes, or water velocities (Spearman, all p>0.192). Our result highlighted specific dissimilarities between males Shelter stability is correlated with CL/TL and ChL/TL and females. Thus, all males expressed an allometric gaining 76 M.I. Groza et al. weight with increase in total length. This boost in male Average chelae size is very similar between individuals weight appears to be the outcome of their accelerated chelae from Aninoasa and Valea Satului, with specific differences development after maturity (Pratten 1980, Grandjean et al. among sexes. However, individuals from Valea Aușoarelor 1997, Endrizzi et al. 2013). Simultaneously, females devel- developed shorter chelae with smaller differences between oped a more isometric variation within individual sizes, males and females. One possible explanation could be shel- with a more balanced weight/length ratio. Similar differ- ter availability. Larger number of more stabile shelters (Fig. ences in weight-length variation between sexes were also 5) mean lower exposure to predators (Vorburger & Ribi underlined in other crayfish populations (e.g Steissl and 1999, Streissl & Hödl 2002b) and probably less time spent by Hodl 2002a, Ďurıš et al. 2006, Endrizzi et al. 2013, Anderson crayfish outside the shelter hunting for prey (Buck et al. & Simon 2015). Both males and females from Aninoasa and 2003). Consequently, individuals could lower energy ex- Valea Aușoarelor expressed an isometric growth while cray- pense by developing smaller chelae. Moreover, smaller che- fish from Valea Satului show a positive allometric develop- lae are advantageous in increasing escape swimming speeds ment with higher gain in weight with increase in length of (Wilson et al. 2009). Another explanation could be the avail- individuals. This is the result of a superior gain in weight of ability of smaller prey. Studies on Carcinus maenas (Edgell & females in Valea Satului that could be influenced by a di- Rochette 2009, Wilcox & Rochette 2015) and Pachygrapsus verse number of factors like: water temperatures, calcium marmoratus (Ferreira Silva et al. 2009) crabs showed positive concentration, availability and quality of food, population changes in the crushing claw morphology determined by density or genetic background (Maguire and Klobucar 2011). prey toughness and size. Further, molluscivorous crabs pre- Generally, shape and size of chelae and cephalothorax fer smaller prey where abundant maximizing energy intake are the most important factors in differentiating intraspecific or minimizing handling time or reduce the risk of claw dam- crayfish populations (Sint et al. 2005, Sint et al. 2007). These age (Seed & Hughes 1995). Also, shorter crushing claw in traits seem to be modeled by several environmental compo- crabs constitutes an advantage in feeding with harder prey nents. For example, Haddaway et al. (2012) found that the (Yamada et al. 2010). However, further investigation on widening of the cephalothorax on Austropotamobius pallipes is crayfish is needed to correctly assume this supposition. linked to lentic habitats or lower flow streams because of limited oxygen. This is also consistent with other findings In-situ individuals’ distribution (Chambers et al. 1979, Sint et al. 2005). On the same note, Crayfish occupied a large array of suitable shelters in the chelae size and shape appear to be affected by variation in streams. However, the distribution of individuals was posi- flow velocity, predator pressure, competition for mate, food tively correlated to their size, with larger and stronger indi- and shelter within species and interactions between species viduals occurring in the most stable shelters along the (Haddaway et al. 2012, Hudina et al. 2012). We also found stream and in the higher elevation segments of the habitat. specific variation in cephalothorax and chelae lengths be- The occupancy of the refuges was most probably consistent tween populations. CL/TL ratio is significantly higher in with social position of individuals as crayfish develop strong Aninoasa, succeeded by Valea Aușoarelor and Valea Satului. size-based hierarchies across the population (e.g., Goess- The length of cephalothorax follows, thus the altitude gradi- mann et al. 2000, Callaghan et al. 2012). Simultaneously, lar- ent, increasing with elevation (r=0.250). Similarly to our ger individuals are more capable of living in faster water findings, Vamosi et al. (2007) linked body size in Dytiscidae flows correlated with the higher altitudes (Pârvulescu et al. with altitudes (when altitudes were lower than 1200m). The 2016). Similarly, the length of the cefalothorax appears to be pattern of increase in altitudes is positively correlated to wa- an important adaptation for higher velocities as we found a ter velocity and oxygen availability (Pârvulescu & Zaharia positive correlation between its size and altitude and water 2013, Endrizzi et al. 2013). Thus, a raise in oxygen concentra- speed. All individuals seem to choose shelters that are position should be also linked to the increase in body size for our tioned sideways from faster water flows. More than 60% of individuals. However, our measurements revealed very the crayfish were attracted by slower running or almost similar concentrations of oxygen in all streams despite the stagnant waters (water velocity smaller than 5 cm/s) in front difference in altitude between sites. Thereby, the increase in of the shelter, independent of the adjacent maximum water body size with the altitude could be a result of the difference speed. Similar results were obtained by Nakata et al. (2003), in oxygen intake by individuals (not oxygen availability), a which revealed a preference of Cambaroides japonicus for low process moderated by temperature, salinity and pressure water speeds (5cm/s) near and inside the shelter. Pöckl & (Peck & Chapelle 2003). Chapelle & Peck 1999 also found an Streissl (2005) also found 40% of A. torrentium in waters be- increase in dissolved oxygen in the haemolymph of amphi- tween 0 and 5 cm/s. Nonetheless, almost 40% of all our in- pods consistent with a decrease in temperature and salinity. dividuals tolerated higher water flows at higher altitudes, As these factors weren’t the focus of out study, a future in- both in front of the gallery and in nearby proximity to the depth analysis is needed to confirm this hypothesis. Perry et shelter. Yet, even for this larger crayfish, the average speed al. (2013) found a negative relation between mean water of the water at the entrance did not exceed 72% of the maxi- speed and Orconectes rusticus body size. Also, studies on fish mum adjacent current. Thus, our results show that individu- revealed species dependent changes determined by different als are choosing the most durable shelters that are posi- water velocities. Salaria fluviatilis developed a more slender tioned in a partially milder section of the stream width as to body shape and longer fins (Laporte et al. 2016). Pakkasmaa protect them from the highest nearby current. Pârvulescu et & Piironen (2000) found a similar pattern for brown trout ju- al. (2016) associated the presence of A. torrentium in faster veniles and a different pattern for salmon juveniles. Salmon flows areas with the existence of more durable shelters that developed a more robust body with increased water flow. offer protection in case of floods. 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