# Origin and Genetic Diversity of Barbatula (Cypriniformes: Nemacheilidae) in Italy

## TL;DR

This study identifies three distinct stone loach species in Italy using genetic data, suggesting they should be treated as separate conservation units.

## Contribution

The study reveals three genetically distinct Barbatula species in Italy, challenging previous assumptions about their taxonomy and conservation status.

## Key findings

- Three highly divergent mitochondrial lineages were identified in Italian Barbatula populations.
- The lineages correspond to three species: Barbatula pironae, Barbatula fluvicola, and Barbatula aff. barbatula.
- The findings suggest these species are native to Italy and require updated conservation assessments.

## Abstract

Recent morphological and molecular studies suggested the existence of several undescribed species within the genus Barbatula. The stone loach (
Barbatula barbatula
) is considered, according to the Italian Red List, as native in Northern Italy and classified as vulnerable (VU), having a limited and fragmented distribution from Lombardy to Friuli‐Venezia Giulia regions. In the present study, 248 specimens of Barbatula sp., collected from 17 sampling sites in Italy—spanning its entire known distribution area—and from one site in Austria, were analysed by sequencing the Cytochrome C Oxidase I (COI) and the Cytochrome B (CytB) mitochondrial regions. Sequencing results were then compared with reference samples from the literature. Three highly divergent mitochondrial lineages were observed in Italian populations, which can be associated with three different species: Barbatula pironae in Friuli‐Venezia Giulia, Barbatula fluvicola in Trentino‐Alto Adige and Lombardy, and Barbatula aff. barbatula coexisting with the latter in Lombardy. The three species, with the first having a distribution limited to the upper Adriatic area, and the other two having a wider distribution north of the Alps, should therefore be considered as different Management Units. Therefore, the integration of the Italian freshwater fish species checklist and the update of their taxonomy are strongly advised. Our data together with other available evidence suggest that the three species are likely native to Italy, and hence a revision or definition of their conservation status might be needed.

The stone loach (
Barbatula barbatula
Linnaeus, 1758) is considered native in Northern Italy and classified as vulnerable (VU), having a limited and fragmented distribution In the present study, 248 specimens of Barbatula sp., collected from sampling sites spanning its entire known distribution area in Italy, were analysed by sequencing the Cytochrome C Oxidase I (COI) and the Cytochrome B (CytB) mitochondrial regions. Three highly divergent mitochondrial lineages were observed in Italian populations, which can be associated to three different species: Barbatula pironae in Friuli‐Venezia Giulia, Barbatula fluvicola in Trentino‐Alto Adige and Lombardy, and Barbatula aff. barbatula coexisting with the latter in Lombardy.