CONSERVATION OF THE APENNINE YELLOW-BELLIED TOAD BOMBINA VARIEGATA PACHYPUS IN LIGURIA (NW ITALY)

Abstract

The Apennine Yellow-bellied toad, Bombina variegata pachypus is endemic to the Italian peninsula. Land abandonment, chytrid infection and climate change are negatively impacting Yellow-bellied toad populations in many parts of its range. In Liguria, a northwestern administrative Italian region, a regional conservation programme has been implemented to contrast the decline. Breeding sites were restored and created, the size and the sanitary status of populations was monitored and restocking was carried out. In the period 2011-2014, nine artificial breeding sites were created, four semi-natural water bodies and a small breeding facility was also created from which tadpoles were introduced in a newly-created site. In Liguria, Apennine Yellow-bellied toad populations are relatively small, but appear stable and non-infected by Batracochytrium dendrobatidis. Many of the new or restored breeding sites were colonised by the Apennine Yellow-bellied toad or by other amphibians, and a new reproductive population of Bombina variegata pachypus became established after restocking. Overall, these results confirm that the creation of small artificial breeding sites is a very successful strategy for conserving amphibian populations. This strategy is also useful for maintaining traditional rural activities in the Mediterranean landscape.

Full text

Herpetological Facts Journal. 2014, 1. ISSN 2256-0327. Supplement 1: Proceedings of the 2
nd
international
Scientific Conference – Workshop “Research and conservation of European herpetofauna and its environment:
Bombina bombina, Emys orbicularis, and Coronella austriaca”
97
CONSERVATION OF THE APENNINE YELLOW-BELLIED TOAD BOMBINA VARIEGATA
PACHYPUS IN LIGURIA (NW ITALY)
Sebastiano Salvidio
1
, Fabrizio Oneto
1
, Dario Ottonello
2
, Luca Braida
1
, Caterina
Ferravante
3
, Elena Grasselli
1
, Giulia Vecchione
1
, Stefano Canessa
4
, Attilio Arillo
1
,
Massimiliano Cardelli
3
1
DISTAV, Università di Genova, Genova, Italy. salvidio@dipteris.unige.it
2
DAIS Università Cà Foscari, Venezia, Italy.
3
Parco Montemarcello-Magra, Sarzana, Italy.
4
University of Melbourne, Australia.
ABSTRACT
The Apennine Yellow-bellied toad, Bombina variegata pachypus is endemic to the
Italian peninsula. Land abandonment, chytrid infection and climate change are
negatively impacting Yellow-bellied toad populations in many parts of its range. In
Liguria, a north-western administrative Italian region, a regional conservation
programme has been implemented to contrast the decline. Breeding sites were
restored and created, the size and the sanitary status of populations was
monitored and restocking was carried out. In the period 2011-2014, nine artificial
breeding sites were created, four semi-natural water bodies and a small breeding
facility was also created from which tadpoles were introduced in a newly-created
site. In Liguria, Apennine Yellow-bellied toad populations are relatively small, but
appear stable and non-infected by Batracochytrium dendrobatidis. Many of the
new or restored breeding sites were colonised by the Apennine Yellow-bellied toad
or by other amphibians, and a new reproductive population of Bombina variegata
pachypus became established after restocking. Overall, these results confirm that
the creation of small artificial breeding sites is a very successful strategy for
conserving amphibian populations. This strategy is also useful for maintaining
traditional rural activities in the Mediterranean landscape.
Keywords: artificial tanks, Batracochytrium dendrobatidis, Bombina, rural
landscape
INTRODUCTION
The Apennine Yellow-bellied toad, Bombina variegata pachypus (Bonaparte, 1838), is an
amphibian endemic to the Italian peninsula. The taxonomic status of the Apennine
populations of Yellow-bellied toad is still debated, because some authors consider them

Herpetological Facts Journal. 2014, 1. ISSN 2256-0327. Supplement 1: Proceedings of the 2
nd
international
Scientific Conference – Workshop “Research and conservation of European herpetofauna and its environment:
Bombina bombina, Emys orbicularis, and Coronella austriaca”
98
as belonging to a subspecies of the Yellow-bellied toad Bombina variegata (Linnaeus,
1758) (Canessa et al., 2013b), while others treat them as full species (Lanza et al., 2007;
Canestrelli et al., 2007). In any case, the Apennine Yellow-bellied toad distribution is
confined to the Apennine Mountains and ranges from Liguria in the North to Calabria in
the South (Lanza et al. 2007) (Figure 1).
Figure 1. Range of the Italian endemic toad Bombina variegata pachypus.
In Liguria (NW Italy), the Apennine Yellow-bellied toad breeds in small temporary streams
and in small standing natural and human-made water bodies (Canessa et al., 2013b). In
recent years, the northern populations of this species are declining because of habitat
loss, abandonment of traditional agricultural activities (Canessa et al. 2013b) and
diffusion of the chytrid fungus pathogen Batracochytrium dendrobatidis (Bd) (Canestrelli
et al., 2013). To reverse this negative trend, an integrated regional conservation project,
starting in 2011, has been coordinated by the Parco di Montemarcello-Magra and the
University of Genova (Arillo et al., 2013). This programme aimed to locally improve the
conservation status of Apennine Yellow-bellied toad populations and at the same time to
promote the traditional farming activities that, in the rural landscape of Liguria, play an
important role in maintaining the diversity of amphibian populations breeding sites
(Canessa et al., 2013b; Romano et al., 2013). The preliminary results concerning
population size estimation and Bd analyses have been published by Canessa et al.
(2013a). Therefore, in this study we will report only about the results concerning the Bd
screening for 2013, the colonisation of the newly-created artificial or semi-natural
habitats and the restocking activities.

Herpetological Facts Journal. 2014, 1. ISSN 2256-0327. Supplement 1: Proceedings of the 2
nd
international
Scientific Conference – Workshop “Research and conservation of European herpetofauna and its environment:
Bombina bombina, Emys orbicularis, and Coronella austriaca”
99
MATERIALS AND METHODS
Chytridiomycosis detection. All the main populations known in Liguria (Canessa et al.,
2013a) have been screened for the pathogen Batracochytrium dendrobatidis (Bd) by
means of PCR assay. Yellow bellied toads were swabbed in the field at elast 30 times with
cottone sterile swabs that were conserved at 4° C until DAN esxtraction. The qPCR
method used for Bd detection, is a modification of the standard TaqMan qPCR technique
(Boyle et al., 2004), based on SYBR Green chemistry with slightly modified primers
(Grasselli et al., unpublished data). This method has been fully validated by a comparison
with the Boyle's method (Boyle et al., 2004) and by the quantification of Bd infection on
amphibian samples with known Bd prevalence (Grasselli et al., unpublished data).
Habitat creation and restocking. During the project nine traditional stone tanks were
built, while four standing water bodies were completely restored. All these sites fall
within the historical range of the Apennine Yellow-bellied toad. These new habitats were
created in private land in accordance with landowners that were willing to use them for
irrigation and farming activities. The landowners signed an agreement with the regional
park of Montemarcello Magra, accepting to take care and manage the sites for ten years.
The artificial tanks were made in concrete and local stones and had a capacity of 2-3 m
3
.
The water input was granted by a natural spring, thus incteasing the possibility of a
permanent hydroperiod. In addition, an escape ramp assuring the exit of amphibians
from the tank was always built (Figure 2).
Figure 2. A) artificial tank in construction; B) close up of an escape ramp (arrow); C and D)
two artificial tanks integrated in the rural lanscape.

Herpetological Facts Journal. 2014, 1. ISSN 2256-0327. Supplement 1: Proceedings of the 2
nd
international
Scientific Conference – Workshop “Research and conservation of European herpetofauna and its environment:
Bombina bombina, Emys orbicularis, and Coronella austriaca”
100
The semi-natural ponds that were restored were already used as breeding sites by
amphibians but they were partially excavated to increase their functionality (Figure 3).
Usually they were already used as breeding sites by amphibians.
Figure 3. A small pond during and at the end of restoration.
The breeding facility was also built in private land and is about 9 x 3 m in size (Figure 4). It
contains three small PVC ponds with capacity of 150 l and hosts only a small number of
breeding adults (2-3 pairs) and it is used also for education with primary school students.
Since 2011, tadpoles born in this structure are introduced in one of the newly-created
tanks located within a SCI where the species was once present but went recently extinct.
RESULTS
During the 2013 survey, 100 swabs of B. v. pachypus were analysed for Bd by qPCR
analysis and all of them were negative, confirming the previous results obtained by
Canessa et al. (2013a) that sampled the same populations in 2011 and 212 (Table 1).

Herpetological Facts Journal. 2014, 1. ISSN 2256-0327. Supplement 1: Proceedings of the 2
nd
international
Scientific Conference – Workshop “Research and conservation of European herpetofauna and its environment:
Bombina bombina, Emys orbicularis, and Coronella austriaca”
101
Figure 4. The breeding facility used for raising Yellow-bellied toad tadpoles.
Table 1. Number of Yellow-bellied toads analysed Bd (number infected). Data for 2011
and 2012 form Canessa et al. (2013a).
Site
2011
2012
2013
Pav
4(0)
-
-
Lor1
17(0)
56(0)
48(0)
Lor2
21(0)
43(0)
8(0)
LorP
-
-
2(0)
Pav
17(0)
7(0)
6(0)
Pin
13(0)
35(0)
18(0)
Tev
1(0)
-
-
Moc
-
-
12(0)
LMa
-
-
1(0)
Almost all the new stone tanks were naturally colonised by amphibians, in four cases by
individuals of Bombina v. pachypus, that in three of them established breeding
populations The other species that were observed breeding in the newly-created tanks,
were the Italian crested newt (Triturus carnifex), the Alpine newt (Mesotriton alpestris)
and the Apennine frog (Rana italica). Concerning restocking, since 2011 about 100 large
tadpoles of Bombina v. pachypus were introduced in the newly-created site (Agn in Table
2), where in the spring 2014 they began to breed successfully.

Herpetological Facts Journal. 2014, 1. ISSN 2256-0327. Supplement 1: Proceedings of the 2
nd
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Scientific Conference – Workshop “Research and conservation of European herpetofauna and its environment:
Bombina bombina, Emys orbicularis, and Coronella austriaca”
102
Table 2. Amphibian populations and agricoltual use of the artificial tanks.
Artificial tank
Amphibians present
Farming use
LMa
M. alpestris, T. carnifex
cattle watering
Moc
B. v. pachypus
.
PerS
B. v. pachypus, M. alpestris
cattle watering
PerM
R. italica
-
Pav
B. v. pachypus
irrigation
Lie
B. v. pachypus
horse watering
Seg
M. alpestris
-
Pig
-
-
Agn
B. v. pachypus (restocked), R.
italica
-
DISCUSSION
In the Mediterranean landscape, small seasonal aquatic habitats represent important
freshwater ecosystems, in which rare organisms of conservation interest may be present
(Blondel and Aronson 1999). In this region, small artificial water reservoirs are still used in
traditional irrigation and may become relevant breeding sites for amphibians (Canessa et
al., 2013b; Romano et al., 2014).
The results of the present study strengthen the importance of artificial tanks as breeding
sites for amphibian populations. In fact, the colonisation of many newly-created artificial
tanks was rapid, beginning few months after their construction especially when
amphibian populations were already present in the area. Even the rare and declining
Apennine Yellow-bellied toad was able to successfully colonize these newly-created tanks,
suggesting that one of its main limiting factor in NW Italy is the absence of suitable
breeding sites. In rural lands, artificial water reservoirs are periodically managed by
landowners. They have interest to assure constant water supply and they periodically
clean the tanks to reduce aquatic vegetation and silting.
These management practices reduce the density of invertebrate predators (mainly
dragonfly larvae) and by consequence may increase the survival and matamorphosis rate
of Yellow-bellied toad tadpoles (Canessa et al., 2013b). It is interesting to note that in one
case, the restocking of about 100 large tadpoles was capable to establish in only three
years a new breeding population of the Apennine Yellow-bellied toad. Therefore, the
strategy of creating artificial sites together with the restocking of tadpoles should be
viewed as a practical and effective conservation strategy for this species (Arillo et al.
2013).

Herpetological Facts Journal. 2014, 1. ISSN 2256-0327. Supplement 1: Proceedings of the 2
nd
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Scientific Conference – Workshop “Research and conservation of European herpetofauna and its environment:
Bombina bombina, Emys orbicularis, and Coronella austriaca”
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Finally, we observed a positive attitude of local people towards amphibians, that are in
general perceived as indicators of traditional and extensive farming. Therefore, in
Mediterranean regions of Italy, many human-made water bodies may be important for
maintaining traditional agriculture while providing breeding sites for amphibian
populations at the same time.
ACKNOWLEDGEMENTS
The Italian Ministry of Environment issues capture, breeidng and restocking permits
(number 2010 DPN-2010-0010807).
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nd
international
Scientific Conference – Workshop “Research and conservation of European herpetofauna and its environment:
Bombina bombina, Emys orbicularis, and Coronella austriaca”
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