The Neretva dwarf goby Orsinigobius croaticus (Gobiiformes, Gobionellidae) is an endemic
fish native to the freshwaters of the Adriatic Basin in Croatia and Bosnia and
Herzegovina, a Mediterranean Biodiversity Hotspot. Due to its limited distribution
range, specific karst habitat and endangered status, laboratory studies on reproductive
biology are scarce but crucial. Herein, we investigated the sound production and
acoustic behaviour of the endangered O. croaticus during reproductive intersexual
laboratory encounters, utilising an interdisciplinary approach. We also performed dissections
and micro-computed tomography (μCT) scanning of the pectoral girdle to
explore its potential involvement in sound production. Finally, comparative acoustic
analysis was conducted on sounds produced by previously recorded soniferous sand
gobies to investigate whether acoustic features are species-specific. The endemic
O. croaticus is a soniferous species. Males of this species emit pulsatile sounds composed
of a variable number of short (~15 ms) consecutive pulses when interacting with
females, usually during the pre-spawning phase in the nest, but also during courtship
outside the nest. Pulsatile sounds were low-frequency and short pulse trains (~140 Hz,
<1000 ms). Male visual behaviour rate was higher when co-occurring with sounds
and females entered the male's nest significantly more frequently when sounds were
present. Characteristic body movements accompanied male sound production, such
as head thrust and fin spreading. Furthermore, μCT scans and dissections suggest that
O. croaticus shares certain anatomical similarities of the pectoral girdle (i.e. osseous
elements and arrangement of levator pectoralis muscles) to previously studied sand
gobies that could be involved in sound production. Multivariate comparisons, using
sounds produced by eight soniferous European sand gobies, effectively distinguished
soniferous (and sympatric) species based on their acoustic properties. However, the
discrimination success decreased when temperature-dependent features (sound
duration and pulse repetition rate) were excluded from the analysis. Therefore, we
suggest both spectral and temporal features are important for the acoustic differentiation
of sand gobies