Biogeographia – The Journal of Integrative Biogeography

New distributional the SUMMARY Scarce data are currently available about the distribution of the Mediterranean medicinal leech Hirudo verbana in Italy, and most of the known occurrence localities are based on records collected in the and the first half of the twentieth century, which were not confirmed in the last decades, mostly due to a lack of surveys. Accordingly, the available knowledge on the distribution of the species is far from being updated and representative, although a significant negative trend of H. verbana throughout the country is supposed. The lack of sound distribution data is a significant shortfall, which hinders the implementation of the monitoring of the species as required by the Article 17 of the “Habitats Directive” on the species of Union concern. To provide recent, validated distributional data for the Mediterranean medicinal leech in Italy to be used as baseline data for further surveys and monitoring, we present herein a set of unpublished observations collected in the last decades in peninsular Italy, Sicily, and Sardinia. Moreover, we report observation of H. verbana feeding on amphibians, a feeding habit to date poorly documented for the Mediterranean medicinal leech. Based on both published and novel data, H. verbana is characterised by a large but fragmented distribution in Italy. Therefore, dedicated monitoring programs and the collection of validated occasional observations are highly desirable to get a clearer picture of the real distribution of the species.


INTRODUCTION
The Mediterranean medicinal leech Hirudo verbana Carena, 1820 is the only Italian leech included in the European "Habitats Directive" (EU Directive 92/43/CEE), where it is listed in Annex 5 sub Hirudo medicinalis Linnaeus, 1758 (i.e., the European medicinal leech). In fact, different medicinal leech species were lumped under this binomen when the "Habitats Directive" came into force, until recent morphological and molecular evidence unequivocally proved that they are distinct species (Trontelj and Utevsky 2012;Saglam et al. 2016). Based on available evidence, the native distribution of Hirudo verbana ranges from Spain to Turkey, including eastern European steppe landscapes, the Caucasus and Uzbekistan, and the populations of the species are subdivided into three major clades, i.e., Iberian, Western and Eastern ones, which can be explained by distinct postglacial colonisations from separate refugia (Trontelj and Utevsky 2012;Arias et al. 2021).
In Italy, Hirudo verbana has a scattered distribution, and only few records of the species were reported in the last decades, often sub Hirudo medicinalis (Bazzanti et al. 1996;Fontaneto et al. 1999;Minelli 2006;Utevsky et al. 2010;Sorgi et al. 2011;Lapini et al. 2013;Osella and Pannunzio 2013;Marrone and Canale 2019). The vast majority of the known occurrence localities derive from observations carried out in the nineteenth and the first half of the twentieth century (see references in Minelli 1979in Minelli , 2006. The scarcity of recent records and the lack of confirmation for most of the historical occurrence localities is generally ascribed to "a significantly (or rather, dramatically) negative trend" of the species throughout the country (Minelli 2006), although the lack of targeted surveys of Hirudo verbana in Italy prevented obtaining an adequate picture of its current distribution, status, and trend.
For this reason, we report herein a collection of unpublished occurrence data for Hirudo verbana in Italy, with the aim of providing reliable and geo-referenced baseline data to be used for future studies and as a reference for the monitoring of the species under the provisions of Article 17 of the "Habitats Directive". Moreover, the observation of feeding behaviour on amphibians is here reported and documented.

MATERIALS AND METHODS
The occurrence of the species was occasionally registered in the frame of sampling activities carried out in lentic and lotic water bodies across the Italian peninsula, Sardinia, and Sicily ( Fig. 1, Table 1). Moreover, the two records available on the social network iNaturalist (www.inaturalist.org) at 10 March 2021 were considered reliable and were included in the dataset. Observed Hirudo verbana individuals were identified and photographed in situ. The identification of the collected leeches was based on the most updated available identification keys (Neubert and Nesemann 1999;Trontelj and Utevsky 2005;Kutschera 2012).

RESULTS AND DISCUSSION
Hirudo verbana was found in 20 natural and semi-natural water bodies where the occurrence of the species was previously unknown. Moreover, H. verbana was observed in the pond named Rekikenca (Colludrozza, Trieste), where the species was considered locally extinct by Sket (1999, sub Hirudo medicinalis).
The occurrence of Hirudo verbana in two localities in the central Apennines (i.e., sites 1 and 9, Table 1) and in one locality in Calabria (site 5, Table 1) are particularly noteworthy since they are located far away from any published record of the species (Fig. 1). In addition to these validated observations, further occurrence localities of H. verbana in Calabria ("Monte Caloria", WWF Calabria 2015) and in Lombardy ("Valli del Mincio", Rigoni 2010) are reported sub H. medicinalis; however, considering that the real presence and identity of the reported species could not be verified, these sites are not included in Table 1.
In accordance with the known broad ecological adaptability of the species, H. verbana was observed in different types of water bodies, from temporary ponds to permanent lakes, lateral pools of small rivers, and artificial canals, located at altitudes ranging from 1 to 1,559 m a.s.l. in different microclimates and habitats. The apparent absence of well-defined ecological preferences of the species prevents the application of species distribution statistical modelling for defining the areas of its potential presence.
5 Only a few of the water bodies which are apparently suitable for the presence of the species are occupied by H. verbana; this is in agreement with the observation that sporadic, non-deterministic occurrences appear to be characteristic to the species of the genus Hirudo (Utevsky et al. 2010). Lacking dedicated surveys aimed at the census of Hirudinea in Italy, it is utmost likely that the currently known distribution of the species is still largely incomplete and far from being exhaustive. In this situation, the collection and publication of validated occasional observations of H. verbana is a useful approach for trying to get a map of the real distribution of the species.
Hirudo verbana was observed feeding on amphibians both in Sicily (site 14,  (Boulenger, 1882) and Triturus carnifex Laurenti, 1768 at several dates (Fig. 2). Although other Hirudo species are known to regularly feed on amphibians (e.g., Neubert and Nesemann 1999;Merilä and Sterner 2002), such observations are noteworthy for H. verbana, which is currently considered to use ungulates as the main food resource (Kovalenko and Utevsky 2015), and its habit to feed on amphibians was to date documented only on Bufo bufo (Linnaeus, 1758) (Fontaneto et al. 1999). This feeding behaviour suggests that the presence of ungulates is not a necessary prerequisite for habitat selection and species distribution. 6 Based on currently available data, H. verbana is characterised by a large but fragmented distribution area in Italy (Fig. 1). Moreover, the occurrence of native or introduced populations of H. medicinalis in Italy cannot be ruled out based on the scant literature. An assessment of the identity of the extant populations from the sites reported in the previous literature sub H. medicinalis is highly recommended. The scattered distribution in isolated water bodies, if confirmed by further evidence, might likely negatively affect the long-term persistence of Italian medicinal leeches, exposing them to the risk of local extinction. The implementation of a monitoring plan based on standardized sampling surveys and the collection of occasional, but validated, observations is highly desirable, also for the purpose of fulfilling the requirements of the Article 17 of the "Habitats Directive" on the species of Union concern. Eventually, as already suggested by Utevsky et al. (2010), the assessment of the species and its inclusion in the IUCN Red List of Threatened Species (https://www.iucnredlist.org/) is worthwhile, considering that only Hirudo medicinalis is included up to now as NT -Nearly Threatened (Utevsky et al. 2014).