Groundwater biodiversity patterns in the Lessinian Massif of

Groundwater biodiversity patterns in the LessinianMassif of northern Italy

DIANA M. P. GALASSI , FABIO STOCH, BARBARA FIASCA, TIZIANA DI LORENZO AND

ENRICO GATTONE

Dipartimento di Scienze Ambientali, University of L’Aquila, Italy

SUMMARY

1. The distribution patterns of stygobiotic invertebrates were examined with a stratified

sampling design at 197 sites selected among four hydrogeographic basins in the

Lessinian Massif (northern Italy). The sites were approximately evenly distributed among

four hydrogeological zones: unsaturated and saturated zone of karstic aquifers, and

hyporheic and saturated zone of porous aquifers.

2. Outlying Mean Index (OMI) analysis which assesses deviation of habitat conditions

from reference conditions, was used to evaluate the importance of 14 selected environ-

mental variables in shaping groundwater biodiversity patterns in the region (total of 89

stygobiotic species). The measured variables explained 80% of the variability in the data

set.

3. Sampling sites were distributed along the environmental gradients defined by OMI

analysis. Significant differences were detected between karstic and porous site, as well

as among sites located in the four hydrogeological zones. Differences among the four

hydrogeographic basins were not observed.

4. Ordination of stygobiotic species along the environmental gradients was best explained

by historical variables (mainly Wurmian glaciation and age of the underlying geological

formation), while variables related to hydrogeology (mainly pH, calcium concentration

and habitat fragmentation) influenced species distributions in the hydrogeological zones.

An Environmental Integrity Index and nitrate concentration were significantly correlated

with altitude, but appeared not to play a significant role in determining stygobiotic

biodiversity patterns at the regional scale.

5. Results of the OMI analysis were highly significant for all taxa, suggesting that

stygobiotic species are sensitive to the environmental factors studied. Thirty-five species

showed high habitat specialization (OMI index > 10). These species were usually rare and

endemic to the Lessinian Massif. Most of them were found in a single hydrogeological

zone.

6. Quaternary glaciations appear not to have lowered stygobiotic species richness in the

Lessinian Massif. This may be because of the marginal location of the region with

respect to the Wurmian glacier limit and because of extensive networks of fractures in

the vadose zone of the karst, which may have allowed stygobionts to move deep down

in the aquifers to seek refuge during surface freezing and to recolonise ancestral habitats

after the glaciers retreated.

Keywords: biogeography, ecology, ground water, Italy, stygobionts

Correspondence: Diana M. P. Galassi, Dipartimento di Scienze Ambientali, University of L’Aquila, L’Aquila, Italy.

E-mail: [email protected]

Freshwater Biology (2009) 54, 830–847 doi:10.1111/j.1365-2427.2009.02203.x

830 � 2009 Blackwell Publishing Ltd

Introduction

Despite great strides in recent years, the true dimen-

sion of groundwater biodiversity is largely underes-

timated and the distribution of groundwater species

at various spatial scales remains only sketchily

understood (Gibert & Deharveng, 2002; Culver,

2005). This situation stems from the still highly

fragmentary knowledge of groundwater biodiversity

as a whole, compared to the biodiversity of surface

waters, and specifically from the limited number of

studies that have analysed the partitioning of ground-

water biodiversity at different spatial scales (Gibert

et al., 1994a,b, 2000; Ferreira et al., 2005; Dole-Olivier

et al., 2009b; Martin et al., 2009).

It is widely recognised that both contemporary

ecological events and historical contingencies

resulting from evolutionary processes shape

biodiversity patterns in different environments (Stoch,

1995; Drake et al., 1996; Galassi, 2001; Moritz et al.,

2001; Whittaker, Willis & Field, 2001; Gibert &

Deharveng, 2002; Moritz, 2002; Colwell, Rahbek &

Gotelli, 2004). However, it is far from being

understood how these factors interact to control

present species diversity and distributions and,

consequently, composition of groundwater assem-

blages. Spatial patterns of biodiversity have mainly

been interpreted from an ecological perspective. Only

occasionally has the role of historical events been

taken into account, and in most of those cases, only

narrative explanations have been proposed to

elucidate the role of historical factors in determining

species diversity and distributions (Rundle et al., 2002;

Castellarini et al., 2005; Hahn & Fuchs, 2005;

Martin et al., 2005). Approaches that account for

both past events and current constraints are complex,

however, because of possible interactions among a

range of variables, which may obscure the influence of

individual factors in shaping species assemblages.

This paper examines regional patterns of ground-

water biodiversity from both an ecological and

historical perspective. Its aims are: (i) to determine

the relative importance of various environmental

descriptors of species distribution patterns in a region;

(ii) to define habitat preferences of groundwater

species and characteristic species assemblages along

environmental gradients and (iii) to identify the main

historical and ecological variables shaping ground-

water biodiversity patterns at different spatial scales

within the region.

Methods

Study area

The study was carried out in the Lessinian Massif,

which is one of six European regions chosen for

assessing groundwater biodiversity in the PASCALIS

project (Gibert, 2001). The region is located in the

southernmost part of the Venetian pre-Alps in northern

Italy (Fig. 1). The rocky massif of 691 km2 emerged in

the Oligocene, 25–24 million years BP (Boccaletti et al.,

1990). It extends southward in divergent finger-like

ridges that reach the alluvial plain of the River Adige

with a mean slope of 10%. The aquifer discharges on

average about 50 m3 s)1: 15 m3 s)1 through alluvial

deposits, 30 m3 s)1 through stream beds and 5 m3 s)1

through a spring system (Montorio). The ground water

flows from north-west to south-east. The main outlets

are located along the boundaries of the alluvial plain of

the River Adige, where limestone comes in contact with

Quaternary alluvial sediments (Patrizi et al., 2001). The

hydrological regime of the Lessinian Massif is mostly

governed by meteorological events. Infiltrating rainfall

tends to flow vertically through the vadose zone of the

karstic massif and reaches the deep saturated aquifers

in a few hours to a maximum of a few days through an

extended network of large and small fractures in the

limestone strata. The drainage network consists of

tributaries of the River Adige, flowing southward along

narrow steep valleys. Streams are often temporary and

predominantly fed by rainfall, to a lesser extent by

ground water.

The geological structure of the massif is dominated

by carbonate rocks of Cretaceous and Jurassic ages in

the northern part. The basement is represented by a

1000-m deep dolomite stratum, covered by several

limestone banks of different origin, reaching a thick-

ness of about 550 m. Eocene limestone is primarily

located in the southern part of the massif, while

outcrops of volcanic rocks, mainly basaltic, are located

in the eastern part. Quaternary alluvial deposits,

covering the limestone basement, fill the smaller

valleys, as well as the broad Adige plain, where

thickness of the alluvial sediments ranges from 100 to

200 m (Patrizi et al., 2001).

Groundwater biodiversity in the Lessinian Massif 831

� 2009 Blackwell Publishing Ltd, Freshwater Biology, 54, 830–847

Sampling

The stratified sampling approach used in the study

area followed the standard protocol of the PASCALIS

project, designed to capture most of the environmen-

tal heterogeneity at different spatial scales (Dole-

Olivier et al., 2009a). Four spatial scales were defined:

(i) region (Lessinian Massif); (ii) hydrogeographic

basin (four basins: Progno di Fumane, 43 km2; Progno

di Valpantena, 154 km2; Vaio di Squaranto, 94 km2;

Alpone-Tramigna, 137 km2); (iii) aquifer type (karstic

and porous) and (iv) hydrogeological zone (unsatu-

rated and saturated zone of karstic aquifers, and

hyporheic and saturated zone of porous aquifers). A

total of 197 sites were approximately evenly distrib-

uted among the four hydrogeological zones in the

four basins (Fig. 1). Caves were sampled in the

unsaturated (vadose) zone of karstic aquifers (Ku),

springs and wells in the saturated zone of karstic

aquifers (Ks), interstitial habitats in the hyporheic zone

of porous aquifers (Ph), and wells in the saturated

zone of porous aquifers (Ps).

Fourteen environmental variables were chosen as

environmental descriptors of stygobiotic species dis-

tribution in the region and measured at each site: (i)

altitude, determined with a geographic positioning

system (GPS III Plus�; Garmin (Europe) Ltd, South-

ampton, U.K.) and verified on topographic maps; (ii)

temperature; (iii) specific conductivity at 25 �C; (iv)

dissolved oxygen concentration and (v) pH, measured

in the field with a multiparametric probe (ECM

Multi�; Dr Lange GmbH, Dusseldorf, Germany); (vi)

calcium concentration, measured by a titrimetric

method with EDTA; (vii) magnesium concentration,

obtained by difference between total hardness and

calcium hardness; (viii) nitrate and (ix) phosphate

Fig. 1 Map of the Lessinian Massif of northern Italy showing the distribution of sampling sites.

832 D. M. P. Galassi et al.


concentrations, measured spectrophotometrically

(HACH DR 2000; Hach Co., Loveland, CO, U.S.A.)

with the cadmium reduction method and the ascorbic

acid method, respectively. Five additional environ-

mental variables were determined at each site: (x)

distance to the Wurmian glacier border on

palaeogeographic maps (Sauro, 1973); (xi) an

Environmental Integrity Index according to the

CORINE Land Cover classification (CLC; CEC,

1994), available at the web site of the European

Environmental Agency (http://www.eea.europa.eu);

(xii) age of the underlying geological formation

(hereafter called geological age); (xiii) degree of

habitat fragmentation and (xix) degree of hydrological

isolation from the surface. The Environmental

Integrity Index was developed by first defining the

extent of the area of interest, assessed in different

ways depending on the hydrogeological features of

each sampling site: (i) for springs, the catchment area;

(ii) for cave streams, the functional catchment area of

the river, and for the vadose habitats, a surface area

corresponding to the extension of the underlying

karstic vadose zone; (iii) for hyporheic sites, a strip 50-

m wide along the river banks from the sampling site

to the source; (iv) for wells, a circular surface (200 m

diameter) around the well. In a second step, the per

cent land surface area occupied by the dominant CLC

category was assessed. The nominal variables no.

xi–xiv were scored according to the criteria described

in Table 1.

Table 1 Basic criteria used for the physiographic description of sampling sites in the Lessinian Massif

Score

Environmental integrity index*

3. Forests and semi-natural areas

3.1. Forests 5

3.2. Shrub and ⁄ or herbaceous vegetation 4

2. Agricultural lands

2.3. Pasture 3

2.4. Heterogeneous agricultural areas 3

2.1. Arable land 2

2.2 Permanent crops 2

1. Artificial surfaces 1

Geological age

Jurassic limestone 4

Cretaceous limestone 3

Eocene limestone 2

Quaternary sediments 1

Habitat fragmentation

Rimstone pool or trickle 4

Saturated karst 3

Phreatic groundwater in unconsolidated sediments or spring 2

Subterranean river or hyporheic habitat 1

Hydrological isolation from the surface**

Porous

Thickness of unsaturated zone Recharge by the river Permeability (m s)1)

>10 m >10)6 3

5–10 m No 10)4 2

<5 m Yes <10)3 1

Karstic

Karstification level Distance to the surface Type of karst

Low >100 m 3

Medium 10–100 m Binary 2

High <10 m Unitary 1

*Nomenclature and scoring follow the standard CORINE land cover classification (CEC, 1994).

**The final score is obtained by the sum of the values assigned to each parameter measured for porous and karstic sites,

respectively.



Faunal samples were taken and treated according to

a standardised protocol (Malard et al., 2002), using a

range of sampling methods and devices. Vadose

waters dropping from cave ceilings were filtered

using a plankton net (60-lm mesh size) and a funnel.

Rimstone pools, trickles and temporary streams in

caves were sampled with small hand pumps or by

sieving sediments through a hand net (60-lm mesh

size). Baited traps were used to collect predatory

amphipods from larger pools and siphons in caves.

Ground water from saturated karstic and porous

aquifers was sampled from wells (3–240 m depth)

equipped with pumps, and by sieving the water

(500 L) at the pump outlet with a hand net (60-lm

mesh size). In some cases, the saturated karst was

sampled using a hand net (60-lm mesh size, 25 cm in

diameter), positioned near the mouths of karstic

springs, by stirring the sediment upstream and

sieving the water immediately downstream of the

disturbed area. Hyporheic habitats were sampled

using a Bou-Rouch pump (Bou & Rouch, 1967) by

extracting and filtering 5 L of interstitial water and

sediments through a hand net (60-lm mesh size). Four

replicate samples were collected at a depth of 50 cm

below the stream bed at each site to cover microhabi-

tat heterogeneity. All faunal samples were preserved

in 4% formaldehyde solution. Invertebrates were

sorted, counted and identified, whenever possible to

species level.

Data analysis

Only stygobionts (i.e. obligate groundwater species)

were retained for statistical analyses. Since faunal data

were not quantitative because of difference in the

sampling methods adopted for different habitats, they

were analysed on the basis of presence or absence at a

given site. Relationships between environmental

variables and the composition of stygobiotic assem-

blages were investigated by means of Outlying Mean

Index (OMI) analysis (Doledec, Chessel & Gimaret-

Carpentier, 2000) using the ADEADE-4 software

(Thioulouse et al., 1997). OMI analysis decomposes

the inertia associated with the distribution of a species

into three terms: (i) the OMI, which measures species

marginality (i.e. habitat deviation from a reference

condition corresponding to a species uniformly

distributed along a given environmental gradient);

(ii) tolerance, which corresponds to the dispersion of

sampling sites containing a species along an environ-

mental gradient (i.e. habitat breadth of the species) and

(iii) residual tolerance, which represents the proportion

of variability in the habitat not explained by the

measured environmental variables (Doledec et al.,

2000). The higher the OMI index of a species, the

higher its habitat specialization. Monte-Carlo permu-

tation tests were used to assess statistical significance

of the marginality of each taxon. OMI analysis was also

used for the ordination of environmental variables,

species and sites, where species are positioned along

an environmental gradient based on their average

marginality.

Results

Environmental gradients

A summary of environmental characteristics at dif-

ferent spatial scales in the Lessinian Massif is given in

Table 2. The basins are all roughly north-to-south

oriented, and are defined by repeated longitudinal

environmental gradients. They share the same under-

lying geological formation of ancient Cretaceous and

Jurassic limestone upstream and recent Quaternary

alluvial deposits downstream. Similarly, the highest

anthropogenic pressures occur in the southernmost

part of all basins, whereas the northern parts charac-

terised by forests and semi-natural areas are much

less affected. Mean values of the Environmental

Integrity Index were very similar among the basins,

except for the Progno di Fumane basin, where most

sites are located in the northern, more natural area of

the Lessinian region. Karstic and porous aquifers

showed some differences in habitat fragmentation,

which was higher in karstic aquifers, especially in the

unsaturated karst (Tables 1 & 2).

Sites belonging to the unsaturated karst were gener-

ally characterised by the lowest temperatures

(mean ± SD: 10.1 ± 2.6 �C) and specific conductivities

(mean ± SD: 384 ± 99 lS cm)1), due to short residence

times in the ground of rainfall infiltrating at high

altitude. Most of these sites also experienced low

anthropogenic disturbance, as reflected in lower

nitrate concentrations (mean ± SD: 11.9 ± 8.3 mg L)1)

compared to saturated porous (mean ± SD: 29.7 ±

18.2 mg L)1) and karstic aquifers (mean ± SD: 20.7 ±

13.5 mg L)1). The hyporheic sites were characterised by

recent Quaternary alluvial deposits, high connectivity



to the surface, the lowest degree of habitat fragmenta-

tion (scores never exceeded the value of 1.0), and

intermediate scores of the Environmental Integrity

Index (mean ± SD: 2.8 ± 1.0). The saturated porous

and karstic aquifers had similar hydrochemistry,

differing from the other hydrogeological zones by

lower oxygen concentrations and higher specific

conductivity and calcium and nitrate concentrations.

The 197 sites sampled were arranged by OMI

analysis along an altitudinal gradient (first axis,

61.7% of total variation) defined by distance to the

Wurmian glacier border, altitude, environmental

integrity and temperature (Fig. 2a). Three additional

variables (magnesium, nitrate and dissolved oxygen

concentration) also contributed to defining the envi-

ronmental gradient, although to a lesser extent. These

ecological and historical factors separated the sites of

three hydrogeological zones along the first OMI axis,

i.e. saturated porous aquifers from saturated and

vadose karstic aquifers (Fig. 2b). The second axis

(18.3% of total variation) was primarily defined by

habitat fragmentation and pH, and secondarily by

calcium concentration and geological age. This gradi-

ent mainly separated hyporheic sites from the sites

located in other hydrogeological zones.

The centroid coordinates of the sites located in each

of the four hydrogeological zones were statistically

different from each other (Student’s t-test, P < 0.0001

after Bonferroni correction for all pair-wise compar-

isons). Conversely, sites grouped by hydrogeographic

basins were not distinguished, indicating no statisti-

cally significant differences between basins (t-test,

P > 0.28).

Species richness

A total of 89 stygobiotic species were identified in the

Lessinian region (Appendix 1). Over one-third of

them (35 species) are new to science. Most of these

were found for the first time during this survey, but a

few were already known from other aquifers in

northern Italy (F. Stoch, unpubl. data). Thirty-six

species have a distribution apparently restricted to the

Lessinian area, i.e. they are strict endemics, and

another 23 species appear to be endemic to small

karstic or alluvial areas in north-eastern Italy. The

highest stygobiotic species richness was found in

Copepoda, followed by Oligochaeta and Amphipoda

(Table 3). Species richness of the unsaturated karstTab

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ðmgL�1Þ

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ðmgL�1Þ

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PO3�4

ðmgL�1Þ

DistancetoW€urmianglacier

borderðkmÞ

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Geologicalage

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isolationfromthesurface

Reg

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197

323

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and hyporheic zone was higher than in the saturated

zones of karstic and porous aquifers, while the

number of species found in the four hydrogeographic

basins was similar (Table 3). Distributional rarity was

exceedingly high: no stygobionts were found in 29

sites (15% of all sites) and 27 species (30% of all

species) occurred in one site only. Even the two most

frequently occurring species, the copepods Nitocrella

psammophila Chappuis and Speocyclops cf. infernus

(Kiefer), were collected in only 27% of the sites.

Species–environment relationships

The first two axes of OMI analysis accounted for 80%

of the marginality of all taxa. The average marginality

of all taxa was highly significant (P < 0.00001; Monte-

Carlo permutation test on the whole data set),

suggesting a strong influence of environmental gra-

dients on the composition of stygobiotic assemblages

(Appendix 2). Thirty-nine species (44%) departed

significantly from a uniform distribution along the

environmental gradient (i.e. they had a statistically

significant marginality: P < 0.05; Monte-Carlo permu-

tation test).

The high deviations (OMI index > 10) shown by 35

species indicate high habitat specialization. Almost all

of these species were found in one of the four

hydrogeological zones only, mostly in unsaturated

karst or in the hyporheic zone (Fig. 2c). Seventeen of

the above 35 species occurred exclusively in the

unsaturated zone of the Lessinian karst (Appendix 2).

Among the species showing a statistically significant

marginality were endemic harpacticoid copepods of

the genus Lessinocamptus (L. caoduroi Stoch, L. pivai

Stoch) and Parastenocaris (Parastenocaris sp. I1), the

endemic bathynellacean Bathynella sp. I1, the endemic

isopod Monolistra (Typhlosphaeroma) berica (Fabiani),

and the amphipod Niphargus forelii Humbert. Eleven

species were recorded exclusively in the hyporheic

zone, but only two of them (the ostracod Fabaeformi-

scandona cf. wegelini Danielopol and the halacaridan

Soldanellonyx visurgis Vietz) showed a statistically

significant marginality. The number of specialised

species found in the saturated zones of porous

(six species) and karstic (two species) aquifers was

low. Among the most ecologically tolerant species

(OMI index < 2) were the gastropod Paladilhliopsis

virei (Locard), the amphipod Niphargus longidactylus

Ruffo and the cyclopoid copepod Diacyclops italianus

(a)

Altitude

pH

Specific conductivity

T Oxygen

Ca2+

Mg2+

NO3-

PO43-

Environmentalintegrity index

Hydrologicalisolation

Distance toglacier

Habitatfragmentation

Geologicalage

0.8

–0.8

1–1

(b) 4

–5

8–6

(c)

Ba1

BaBI1

BaLI1

BaLI2

CaSu

CeSe

CnI1

CnI2

CnI3

CnI4

Cv2P

Di2C

Di2MDiI1

DiI2

DiI3

DiI4

DiItDiPa

DiRu

EcI1

ElEl

ElI1

ElPs

Gi2L

GiI1

GiI2

GrUn

Ha1HaPh

IgCo

Ko1 KoDe

LeCa

LeI1

LeI2

LeIn

LePi

LoWQ

MeI1

MM2C

MMI1

MMSt MTBeNiPs

Np2CNp2F Np2L

NpAq

NpBG

NpFor

NpGa

NpLe

NpLo

NpSiNpTa

NpTr

PaGe

PaI1

PaI2

PaI3

PaI4

PaIt

Ph1

PlVi

PmI1

PmI2

PpIt

PrI1

PrI2

PrI3

Ps2E

PvSp

Rh2D

RhI1

RhI2

SgLa SoCh

Sp2I

Tr2P

TrI1

TroB

3

–3

–3 6

Fig. 2 (a) Ordination of environmental variables (indicated by

arrows) on the space defined by the first two canonical axes

extracted by OMI analysis and explaining 80% of the variability

in the data set; (b) ordination of sampling sites (black cir-

cles = sites in the unsaturated zone of karstic aquifers; grey

circles = sites in the saturated zone of karstic aquifers; black

triangles = sites in the hyporheic zone of porous aquifers; grey

triangles = sites in the saturated zone of porous aquifers); (c)

ordination of stygobiotic species (see Appendix 1 for species

identification codes).



(Kiefer), which are endemic to northern Italy (Appen-

dix 1).

The average position of each species along the first

axis of the environmental gradient defined by OMI

analysis is shown in Fig. 3. The average position of the

two most widely distributed species (large-sized

circles) are located far away from the origin of the

axis, indicating that one of the species (S. cf. infernus)

occurred primarily in the vadose zone of karst and,

less frequently, in the hyporheic zone, while the other

(N. psammophila) mainly occurred in saturated porous

sites. The average positions of the two most widely

distributed species along the second axis of OMI

analysis (Fig. 4) are close to the origin of the axis,

indicating wide ecological tolerance to environmental

variation.

The average positions of the other species are evenly

spaced along the altitudinal gradient. Only two

species, L. caoduroi and Bathynella (Bathynella) sp. I1,

were restricted to high-altitude karstic sites, specifi-

cally to the vadose zone of caves located close to the

Wurmian glacier border. Both species are endemic to

the Lessinian Massif. Two other species, Limnosbaena

sp. and Bathynella (Bathynella) sp. I5, are located at the

opposite side of the environmental gradient. Both were

collected at a single site in a saturated porous aquifer.

Discussion

Environmental descriptors

The groundwater environment in the Lessinian Massif

was described by OMI analysis as a gradient from

high altitude, unsaturated zones of karstic aquifers, to

low altitude saturated karstic and porous zones of

more recent geological age and lower environmental

integrity. A second gradient, mainly defined by pH

and calcium concentration, separated the hyporheic

zone from the saturated zones of both karstic and

porous aquifers. Nonetheless, a clear distinction

between the influence of historical and ecological

variables in separating the four hydrogeological zones

was partially blurred by a systematic spatial segrega-

tion of the hydrogeological zones (arranged in north–

south orientation) in the Lessinian region.

Unsaturated karstic sites developed in ancient Juras-

sic and Cretaceous limestone, and are characterised by

a high degree of habitat fragmentation, resulting in

many microhabitats (Pipan & Culver, 2005; CamachoTa

ble

3S

pec

ies

rich

nes

so

fst

yg

ob

ioti

cin

ver

teb

rate

tax

aat

dif

fere

nt

spat

ial

scal

esin

gro

un

dw

ater

of

the

Les

sin

ian

Mas

sif

Sp

atia

lsc

ale

Sp

ecie

sri

chn

ess

All

tax

aP

oly

chae

taO

lig

och

aeta

Gas

tro

po

da

Cy

clo

po

ida

Har

pac

tico

ida

Ost

raco

da

Iso

po

da

Am

ph

ipo

da

Th

erm

osb

aen

acea

Bat

hy

nel

lace

aA

cari

Reg

ion

891

152

1224

72

121

67

Pro

gn

od

iF

um

ane

bas

in46

07

29

94

17

02

5

Pro

gn

od

iV

alp

ante

na

bas

in42

04

28

122

17

03

3

Vai

od

iS

qu

aran

tob

asin

410

62

716

11

50

21

Alp

on

e–T

ram

ign

ab

asin

511

81

914

41

71

32

Kar

stic

aqu

ifer

s75

110

312

215

113

05

4

Po

rou

saq

uif

ers

650

92

1615

71

51

36

Un

satu

rate

dk

arst

icaq

uif

ers

421

92

312

11

90

31

Sat

ura

ted

kar

stic

aqu

ifer

s33

01

19

94

04

02

3

Hy

po

rhei

czo

ne

440

91

912

50

20

06

Sat

ura

ted

po

rou

saq

uif

ers

210

01

73

21

31

30



et al., 2006) with different degrees of isolation (Pipan,

Christman & Culver, 2006; Pipan & Culver, 2007).

Hyporheic waters were characterised by the lowest

calcium concentrations measured in this study. Water

flowing through sandy and silty sediments of the

hyporheic zones may account for the relatively low

calcium concentrations compared to ground water of

limestone or gravel aquifers (Bakalowicz, 1994).

Separation of saturated porous and karstic sites

along the axes of OMI analysis is less sharp,

albeit statistically significant, because both are

located at low altitude, in areas with low scores

of the Environmental Integrity Index and high

nitrate concentrations. Consequently, environmental

contrasts among sites in saturated aquifers were

low.

(a)

(c) (b)

Fig. 3 First axis extracted by the OMI analysis of stygobiotic assemblages, which explains 61.7% of the variability in the data set:

(a) Environmental variables arranged along the first axis using their canonical weights (environmental gradient); (b) ordination of

the 89 stygobiotic species along the first axis using their factorial scores; (c) distribution of species along the environmental gradient as

a function of their weighted average position along site scores. Small grey circles represent sites where a species occurs, dark grey

circles the centroid (mean position) of a given species along the environmental gradient. Size of the circles is proportional to the total

occurrence of a given species. Horizontal bars represent standard deviations. See Appendix 1 for species identification codes.



Phosphate concentration was not an important

environmental factor in the ordination of sites by

OMI analysis, and the contribution of NO3) was also

weak, albeit statistically significant. This suggests that

groundwater pollution by nutrients plays a minor role

in defining environmental gradients of groundwater

habitats in the region, as was also observed by Hahn

& Fuchs (2005), Paran et al. (2005), Dole-Olivier et al.

(2009b) and Martin et al. (2009).

Habitat preferences of stygobionts

The stygobiotic species collected in ground water of

the Lessinian Massif showing low tolerance (i.e. a

(a)

(b)

(c)

Fig. 4 Second axis extracted by OMI analysis of stygobiotic assemblages, which explains 18.3% of the variability in the data set.

See caption of Fig. 3 for details.



high OMI index) were also rare in a geographical

sense, showing low frequencies of occurrence, which

frequently results in a restricted geographic distribu-

tion as well. This is the case for species found almost

exclusively in the vadose zone of karst and that are

endemic to the Lessinian Massif. Some of these rare

species are new to science, such as Bathynella (Bathy-

nella) sp. I1, Bathynella (Lombardobathynella) sp. I2,

Parastenocaris sp. I4, Paramorariopsis sp. I1 and three

species of the genus Lessinocamptus. Larger stygobiotic

species in the region have a wider geographical

distribution in the Italian Pre-Alpine or Alpine area,

such as the isopod M. (Typhlosphaeroma) berica and

the amphipods Niphargus sp. gr. aquilex Schiodte,

N. lessiniensis Stoch, N. galvagnii Ruffo, and N. forelii

Humbert (Ruffo & Stoch, 2006).

Some species with restricted ecological tolerance

were found exclusively in the hyporheic habitat, e.g.

most species of the oligochaete genera Rhyacodrilus

and Pristina (new to science and strict endemics to

the Lessinian Massif), the ostracod F. cf. wegelini, and

several copepod species of the genera Diacyclops and

Parastenocaris. Such species often occurred only at a

single site. The water mites Kongsbergia dentata

Walter, Stygomomonia latipes Szalay and S. visurgis

Vietz were mostly found in the same environment,

probably because of their strict association with

epigean aquatic insects for phoresy (i.e. attachment

to other animals for transportation) (Davids et al.,

2006).

In deep saturated aquifers, most species showed

wide ecological tolerance, which is also reflected

in their wide geographical distribution [e.g. Elaphoid-

ella elaphoides (Chappuis), N. psammophila, Para-

pseudoleptomesochra italica Pesce & Petkovski,

Niphargus transitivus Sket]. A unique assemblage of

ancient marine origin was found at one site. It

included two Malacostraca [a Limnosbaena species of

the order Thermosbaenacea and the isopod Monolistra

(Monolistra) sp. I1], together with an undescribed

genus of the harpacticoid family Ectinosomatidae.

Local factors may account for this assemblage

composition. This fauna may have survived in a very

restricted part of the Alpone-Tramigna basin in

connate waters, also named fossil aquifers, i.e. con-

fined aquifers in which trapped water has a different

age than the geological formation (Fetter, 1994).

The most tolerant species, displaying a very low

OMI index, had a wide geographical distribution, as

also found by Dole-Olivier et al. (2009b) in a parallel

study in the French Jura. The highest tolerance indices

were found for (i) the gastropod P. virei, the amphi-

pod N. longidactylus, and the cyclopoid copepod D.

italianus, which are restricted to north-eastern Italy

(Appendix 1); (ii) the harpacticoid Moraria (Moraria)

stankovitchi Chappuis, which is widely distributed in

eastern Europe (Petkovski & Brancelj, 1985) and (iii)

the cyclopoid Graeteriella (Graeteriella) unisetigera

(Graeter), which has a wide geographical range across

Europe (Fiers & Ghenne, 2000; Galassi, 2001) and is

one of the most common species in the French Jura

(Dole-Olivier et al., 2009b) and in the Walloon karst of

Belgium (Martin et al., 2009). These species were

collected from different groundwater habitats, springs

and hyporheic sites (Botosaneanu, 1986). Slight niche

shifts among locations together with greater dispersal

potential may explain the wide ecological and bio-

geographical distribution of these tolerant species

(indicated by their low OMI index), as observed for

some copepod species from both ground water

(Galassi, 2001) and surface water (Rundle et al., 2002).

Biodiversity patterns

The Lessinian Massif showed a higher stygobiotic

species richness (89 species) than all but one of the

other European regions investigated in the PASCALIS

project with identical methods. Only the Krim Massif

in Slovenia had more species, a total of 92 (Dole-

Olivier et al., 2009a). This high species richness in

Lessinian ground water is paralleled by a high

number of endemic species, 59 in total, which is equal

to as much as 66% of the total species richness.

Outlying Mean Index analysis indicates that both

historical and ecological variables are influential in

determining species diversity patterns in the four

hydrogeological zones studied. The historical compo-

nent, mostly defined by geological age of the aquifer,

appears to be important in increasing both total

species richness and degree of endemism. This is

especially true in the ancient unsaturated karst, where

habitats are highly fragmented, thus favouring long-

term vicariant events, multiple disjunct refugia and

hence speciation. Relatively high species richness was

also found in the hyporheic habitat, as also noted by

Paran et al. (2005) for a shallow aquifer in France. The

richness level of 44 species in the Lessinian Massif is

comparable to that found in the unsaturated karst of



the region (42 species). In contrast, the saturated

porous and karstic aquifers showed relatively low

species richness, probably reflecting low microhabitat

diversity and, though less clearly, anthropogenic

impacts. Even though the Environmental Integrity

Index and nitrate concentration showed statistically

significant correlations with altitude in the Lessinian

Massif, these variables appear to have played minor

roles in explaining stygobiotic biodiversity patterns at

the regional scale, as also noted for the French Jura

(Dole-Olivier et al., 2009b) and the Walloon karst in

Belgium (Martin et al., 2009). This conclusion is ten-

tative, however, since the potential impact of human

activities on stygobiotic species richness remains

incompletely understood in general (Hancock, 2002;

Hancock & Boulton, 2005; Lafont et al., 2006).

Separation of stygobiotic assemblages in karstic and

porous aquifers and, to a lesser extent, among hydro-

geological zones in the French Jura was also found by

Castellarini et al. (2005). These authors postulated that

differences in physical habitat structure are the major

source of variation in species richness and composi-

tion between karstic and porous aquifers. Neverthe-

less, connectivity between basins within the region

and distance to the Wurmian glacier limit were

important factors accounting for differences in stygo-

biotic assemblages in their study. This contrasts results

obtained by Martin et al. (2009) who failed to find

environmental gradients distinguishing karstic and

porous aquifers or hydrogeological zones in the

Walloon karst of Belgium. The different result of the

study of Martin et al.’s (2009), compared to those of both

Castellarini et al. (2005) in the French Jura and the

present study, could have resulted from strong

influences of the Weichselian glaciation on the Walloon

karst. Stygobiotic species richness in the Walloon karst

is very low (36 species) and most species are habitat

generalists with wide geographical distributions,

suggesting that aquifers in that region were recolonised

after the glaciers retreated (Martin et al., 2009).

In the Lessinian ground water, Quaternary glacia-

tions, although important in shaping stygobiotic spe-

cies distribution, appear not to have strongly affected

species richness. Only a few sites in the unsaturated

karst were covered by the Wurmian glaciers, and they

had high species richness. A similar situation has been

observed by Stoch (2000) for the vadose caves of

Trentino in northern Italy, by Rouch (1986) for different

karstic systems in southern France, and by Holsinger,

Mort & Recklies (1983) for groundwater systems in

Canada, all areas that were ice-covered during the

Pleistocene. Plausible explanations of high stygobiotic

species richness in the Lessinian Massif caves are their

marginal location with respect to the Wurmian glacier

limit and the highly fractured nature of the karst.

Extensive networks of fractures probably allowed

stygobionts to move deep down in the aquifers to seek

refuge during surface freezing and to recolonise the

ancestral habitats after the glaciers retreated.

Results of the present study suggest that the ancient

geological age of aquifers in the Lessinian Massif, high

habitat fragmentation, and isolation of microhabitats

may all have promoted speciation by vicariance,

especially in the vadose zone. In particular, the ancient

age of the Lessinian karst may have favoured persis-

tence of many endemics, most of which are phyloge-

netic relicts (i.e. unique remnants of formerly

diversified taxonomic groups) or distributional relicts

(i.e. taxa with close relatives traceable in disjunct

geographical areas) (Holsinger, 1988; Humphreys,

2000). This is the case for the four species of

Lessinocamptus, a harpacticoid genus known only from

unsaturated karstic sites in eastern Italy (Stoch, 2006),

and for some isopod and amphipod species of the

genera Monolistra and Niphargus. Conversely, high

species richness in the hyporheic habitat may be due

to spatial variability in environmental conditions and

speciation promoted by niche differentiation (Stoch,

1995; Galassi, Huys & Reid, 2009), as suggested by the

co-occurrence of several, closely related species in the

same habitat. This scenario could apply to several

copepod species of the genera Diacyclops and

Parastenocaris.

Acknowledgments

We thank Paola D’Ambrosio (L’Aquila), Gianfranco

Tomasin (Gorizia) and Enrico Mezzanotte (Verona)

who greatly contributed to the sampling. The Adige

River Basin Authority (Trento), the ARPA Verona, the

Museo Civico di Storia Naturale of Verona and the

speleological associations in Verona provided facili-

ties for the field work and hydrogeological maps. A.

Camacho (Syncarida), P. Marmonier (Ostracoda), B.

Sambugar (Oligochaeta) and A. Di Sabatino (Acari)

are greatly acknowledged for species identification.

We are also much indebted to J. Gibert, D. Culver and

M. Gessner for useful comments on the final draft of



the manuscript. This study was supported by the

PASCALIS project funded by the European Commis-

sion (contract no. EVK2-CT-2001-00121).

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(Manuscript accepted 11 February 2009)

Appendix 1 List of stygobiotic species collected from ground water in the Lessinian Massif. Two asterisks (**)

indicate species strictly endemic to the Lessinian Massif; one asterisk (*) indicates species endemic to larger

areas of north-eastern Italy

Taxonomic group Endemism Species Identification Code

Polychaeta Troglochaetus beranecki Delachaux TroB

Oligochaeta Cernosvitoviella cf. parviseta Gadzinska Cv2P

** Gianius sp. I1 GiI1

** Gianius sp. I2 GiI2

Gianius cf. labouichensis (Rodriguez & Giani) Gi2L

Haber sp. Ha1

* Parvidrilus spelaeus Martınez-Ansemil, Sambugar & Giani PvSp

Phallodrilinae indet. Ph1

** Pristina sp. I1 PrI1



* Rhyacodrilus cf. dolcei Martınez-Ansemil, Sambugar & Giani, Rh2D

** Rhyacodrilus sp. I1 RhI1

** Rhyacodrilus sp. I2 RhI2

** Trichodrilus sp. I1 TrI1

Trichodrilus cf. pragensis Vejdovsky Tr2P

Gastropoda * Iglica concii (Allegretti) IgCo

* Paladilhiopsis virei (Locard) PlVi

Cyclopoida * Diacyclops cf. maggii Pesce & Galassi Di2M

Diacyclops cf. clandestinus (Kiefer) Di2C

Diacyclops italianus (Kiefer) DiIt

Diacyclops paolae Pesce & Galassi DiPa

* Diacyclops ruffoi Kiefer DiRu

* Diacyclops sp. I1 DiI1



** Diacyclops sp. I4 DiI4

Graeteriella (Graeteriella) unisetigera (Graeter) GrUn

* Speocyclops cf. infernus (Kiefer) Sp2I

** Speocyclops sp. I1 SpI1

Harpacticoida ** Bryocamptus sp. I1 BrI1

Ceuthonectes serbicus Chappuis CeSe

** Ectinosomatidae gen. I1 sp. I1 EcI1

Elaphoidella elaphoides (Chappuis) ElEl

Elaphoidella phreatica (Chappuis) ElPh

* Elaphoidella pseudophreatica (Chappuis) ElPs

* Elaphoidella sp. I1 ElI1

** pivai Lessinocamptus caoduroi Stoch LeCa

** Lessinocamptus insoletus (Chappuis) LeIn

** Lessinocamptus pivai Stoch LePi

** Lessinocamptus sp. I1 LeI1

** Lessinocamptus sp. I2 LeI2

* Moraria (Moraria) sp. I1 MMI1

Moraria (Moraria) stankovitchi Chappuis MMSt

* Nitocrella psammophila Chappuis NiPs



Appendix 1 (Continued)

Taxonomic group Endemism Species Identification Code

Parapseudoleptomesochra italica Pesce & Petkovski PpIt

** Paramorariopsis sp. I1 PmI1

** Paramorariopsis sp. I2 PmI2

* Parastenocaris gertrudae Kiefer PaGe

Parastenocaris italica Chappuis PaIt

** Parastenocaris sp. I1 PaI1




Ostracoda ** Candoninae gen. I2 sp. I1 CnI1

** Candoninae gen. I2 sp. I2 CnI2



Cavernocypris subterranea (Wolf) CaSu

Fabaeformiscandona cf. wegelini Danielopol Fa2W

Pseudocandona cf. eremita (Vejdovsky) Ps2E

Isopoda * Monolistra (Typhlosphaeroma) berica (Fabiani) MTBe

** Monolistra (Monolistra) sp. I1 MM2C

Amphipoda * Niphargus sp. gr. aquilex Schiodte NpAq

* Niphargus bajuvaricus grandii Ruffo NpBG

** Niphargus cf. costozzae Schellenberg Np2C

Niphargus forelii Humbert NpFor

Niphargus cf. forelii Humbert Np2F

* Niphargus galvagnii Ruffo NpGa

** Niphargus lessiniensis Stoch NpLe

** Niphargus cf. lessiniensis Stoch Np2L

* Niphargus longidactylus Ruffo NpLo

* Niphargus similis G. Karaman & Ruffo NpSi

* Niphargus tamaninii Ruffo NpTa

* Niphargus transitivus Sket NpTr

Thermosbaenacea Limnosbaena sp. Th1

Bathynellacea Bathynella sp. Ba1

** Bathynella (Bathynella) sp. I1 BaBI1

** Bathynella (Bathynella) sp. I5 BaBI5

** Bathynella (Lombardobathynella) sp. I1 BaLI1

** Bathynella (Lombardobathynella) sp. I2 BaLI2

** Meridiobathynella sp. I1 MeI1

Acari Stygomomonia latipes Szalay SgLa

Kongsbergia dentata Walter KoDe

Kongsbergia sp. Ko1

Lobohalacarus weberi quadriporus (Walter) LoWQ

Soldanellonyx visurgis Vietz SoVi

Soldanellonyx chappuisi Walter SoCh

Halacarellus phreaticus Petrova HaPh



Appendix 2 Results of the OMI analysis based on OMI index and its variance terms. OMI = Outlying Mean

Index (marginality); T = tolerance (outlying variance); RT = residual tolerance; P = probability value (significance

of the OMI index assessed using Monte-Carlo permutation test). Species are arranged in decreasing order of

marginality. Species with a statistically significant marginality are shown in bold. N = total number of sites where

a taxon occurred. The relative occurrence of species (%) is given within the four hydrogeological zones:

Ku = unsaturated (vadose) zone of karstic aquifers; Ks = saturated zone of karstic aquifers; Ph = hyporheic zone

of porous aquifers; Ps = saturated zone of porous aquifers)

Species OMI T RT P N Ku Ks Ph Ps

Bathynella (Bathynella) sp. I1 38.6 » 0 1.0 0.030 1 1 0 0 0

Lessinocamptus caoduroi 38.6 » 0 1.0 0.030 1 1 0 0 0

Limnosbaena sp. 25.9 0 0 0.054 1 0 0 0 1

Bathynella (Lombardobathynella) sp. I2 22.6 » 0 1.0 0.076 1 1 0 0 0

Bathynella (Bathynella) sp. I5 21.6 0 0 0.092 1 0 0 0 1

Elaphoidella phreatica 20.9 0 0 0.119 1 0 0 1 0

Pristina sp. I3 20.3 » 0 1.0 0.124 1 0 0 1 0

Bathynella sp. 19.8 1.1 5.8 0.020 2 1 0 0 0

Parastenocaris sp. I4 18.2 2.3 2.4 0.027 2 1 0 0 0

Troglochaetus beranecki 17.3 » 0 1.0 0.212 1 1 0 0 0

Niphargus forelii 17.1 » 0 3.8 0.001 4 1 0 0 0

Fabaeformiscandona cf. wegelini 15.9 0.1 1.1 0.013 3 0 0 1 0

Pristina sp. I2 15.5 0 0 0.276 1 1 0 0 0

Niphargus galvagnii 15.3 » 0 1.0 0.306 1 1 0 0 0

Bryocamptus sp. I1 14.5 » 0 1.0 0.316 1 0 0 1 0

Speocyclops sp. I1 12.6 » 0 1.0 0.415 1 0 0 1 0

Gianius sp. I2 12.4 0 0 0.379 1 1 0 0 0

Soldanellonyx visurgis 12.3 » 0 3.2 0.027 3 0 0 1 0

Gianius cf. labouichensis 11.9 0 0 0.411 1 0 0 1 0

Monolistra (Monolistra) sp. I1 11.9 0.3 3.1 0.090 2 0 0 0 1

Lessinocamptus sp. I1 11.8 0 0 0.447 1 1 0 0 0


Niphargus transitivus 11.6 » 0 1.0 0.474 1 0 0 0 1

Bathynella (Lombardobathynella) sp. I1 11.5 0.3 3.4 0.108 2 0 0.5 0 0.5

Elaphoidella elaphoides 11.3 » 0 1.0 0.494 1 0 0 0 1

Candoninae gen. I1 sp. I2 11.1 0 0 0.521 1 0 0 1 0

Lessinocamptus pivai 11.0 0.9 15.9 <0.001 10 1 0 0 0

Rhyacodrilus sp. I1 10.8 » 0 1.0 0.540 1 1 0 0 0

Parapseudoleptomesochra italica 10.5 0.2 2.9 0.049 3 0 1 0 0

Kongsbergia dentata 10.3 » 0 0.8 0.146 2 0 0 1 0

Cavernocypris subterranea 10.2 » 0 2.5 0.167 2 0 0 1 0

Monolistra (Typhlosphaeroma) berica 10.2 0.7 2.5 0.008 5 1 0 0 0

Niphargus sp. gr. aquilex 10.1 0.2 2.2 0.163 2 1 0 0 0

Paramorariopsis sp. I1 10.0 0.1 2.0 0.185 2 1 0 0 0

Trichodrilus sp. I1 10.0 » 0 1.0 0.617 1 1 0 0 0

Moraria (Moraria) sp. I1 9.7 1.7 5.3 <0.001 14 1 0 0 0

Diacyclops ruffoi 9.5 1.4 8.9 <0.001 7 0 0 0 1

Elaphoidella pseudophreatica 9.5 0.6 3.2 0.065 3 1 0 0 0

Paramorariopsis sp. I2 9.5 0 2.0 0.190 2 0 0 1 0

Parastenocaris italica 9.4 1.1 3.9 0.001 7 0 0 1 0

Parastenocaris gertrudae 9.2 0.1 3.4 0.076 3 0 0 1 0

Niphargus cf. forelii 8.4 0 0 0.764 1 1 0 0 0

Niphargus similis 8.3 1.9 6.6 <0.001 8 0.63 0.38 0 0


Halacarellus phreaticus 7.9 0.4 2.2 0.015 5 0 0.20 0.80 0

Rhyacodrilus cf. dolcei 7.9 0 0 0.784 1 0 0 1 0

Rhyacodrilus sp. I2 7.9 0 0 0.784 1 0 0 1 0

Trichodrilus cf. pragensis 7.7 0.7 3.7 0.006 6 1 0 0 0



Appendix 2 (Continued)

Species OMI T RT P N Ku Ks Ph Ps

Kongsbergia sp. 7.7 0.3 1.6 0.316 2 0 0 1 0

Lessinocamptus sp. I2 7.5 1.9 5.5 0.050 4 0.75 0.25 0 0

Niphargus cf. costozzae 7.4 0.2 4.0 0.005 7 1 0 0 0

Pristina sp. I1 7.2 0.1 1.2 0.342 2 0 0 1 0

Diacyclops sp. I4 7.0 » 0 1.0 0.886 1 1 0 0 0

Meridiobathynella sp. I1 6.9 4.1 10.5 0.021 6 0 0.33 0 0.67

Niphargus lessiniensis 6.8 0.8 5.3 <0.001 14 0.86 0.14 0 0

Niphargus cf. lessiniensis 6.8 0.1 3.4 0.066 4 1 0 0 0

Stygomomonia latipes 6.6 » 0 1.0 0.873 1 0 0 1 0

Haber sp. 6.6 1.7 2.9 0.049 5 0.2 0 0.8 0

Niphargus tamaninii 6.4 0 4.1 0.045 5 1 0 0 0

Iglica concii 5.8 0.6 4.4 0.001 13 0.85 0.15 0 0

Parastenocaris sp. I3 5.6 » 0 1.0 0.963 1 0 1 0 0

Soldanellonyx chappuisi 5.2 0.2 4.5 0.285 3 0 0.33 0.67 0

Gianius sp. I1 5.1 1.2 4.7 0.336 3 0.33 0 0.67 0

Lobohalacarus weberi quadriporus 5.1 1.7 4.6 <0.001 18 0 0.11 0.89 0

Pseudocandona cf. eremita 4.9 0.7 5.9 0.618 2 0 0.5 0 0.5

Diacyclops sp. I3 4.8 1.8 4.6 0.022 8 0 0.13 0.13 0.75

Ectinosomatidae gen. I1 sp. I1 4.8 1.8 7.2 0.005 11 0 0.18 0.18 0.64

Ceuthonectes serbicus 4.8 0.5 6.2 <0.001 16 0.63 0.06 0.31 0

Lessinocamptus insoletus 4.7 0.5 4.2 0.015 9 0.56 0.44 0 0

Parvidrilus spelaeus 4.6 1.7 7.8 0.008 9 0.78 0 0.22 0

Speocyclops cf. infernus 4.5 1.7 10.3 <0.001 54 0.63 0.04 0.33 0

Cernosvitoviella cf. parviseta 4.1 » 0 3.6 0.749 2 0 1 0 0

Niphargus bajuvaricus grandii 4.0 0.9 9.6 0.011 10 0 0.1 0.3 0.6

Candoninae gen. I2 sp. I1 4.0 » 0 1.00 0.989 1 0 1 0 0

Elaphoidella sp. I1 3.9 1.3 9.4 0.006 13 0.15 0.23 0.62 0

Candoninae gen. I1 sp. I1 3.8 1.3 7.2 0.313 4 0 0.25 0.5 0.25

Candoninae gen. I2 sp. I2 3.7 2.4 5.7 0.008 11 0.18 0.18 0.64 0

Diacyclops paolae 3.6 2.0 9.1 <0.001 21 0 0.10 0.24 0.67

Diacyclops sp. I2 3.5 1.4 4.7 0.244 5 0 0.2 0.8 0

Diacyclops sp. I1 3.4 1.3 8.5 0.001 20 0 0.3 0.2 0.5

Diacyclops cf. maggii 3.3 0.9 4.2 0.841 2 0 0.5 0 0.5

Diacyclops cf. clandestinus 2.8 1.6 8.4 0.023 12 0 0.08 0.67 0.25

Nitocrella psammophila 2.8 2.0 9.0 <0.001 54 0 0.28 0.24 0.48

Phallodrilinae indet. 2.3 2.3 14.0 0.015 16 0.56 0 0.438 0

Graeteriella (Graeteriella) unisetigera 2.1 3.9 6.9 0.040 15 0 0.27 0.73 0

Moraria (Moraria) stankovitchi 1.5 3.0 5.4 0.745 5 0.4 0.4 0.2 0

Paladilhliopsis virei 1.4 2.8 6.9 0.323 11 0.73 0 0.18 0.09

Niphargus longidactylus 1.1 0.9 9.2 0.945 4 0 0.25 0.5 0.25

Diacyclops italianus 0.6 1.5 12.7 0.369 22 0.45 0.14 0.14 0.27



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Groundwater biodiversity patterns in the Lessinian Massif of