Pomatoschistus marmoratus and Pomatoschistus microps are small sedentary gobies inhabiting the lagoons of European Mediterranean and Atlantic coasts. Along the French Mediterranean coast their respective geographical distribution is not precisely known, in part because they are cryptic species. In this study, 512 gobies of both species were caught as 17 samples in 12 lagoons of the Golfe du Lion on the French Mediterranean coast. They were genotyped at six microsatellite loci and investigated statistically using multidimensional analyses, Bayesian assignment (Structure) and Newhybrids classification. This allowed us to describe the contrasted distribution of each species (P. microps in the east, P. marmoratus in the west), with several exceptions. Neither geographic structure nor isolation by distance was detected among differentiated populations of each species. The suggested mechanism is a deep sedentary behaviour associated with foundations following extinctions. The two species are sympatric or even in syntopy in 5 or 6 sampled lagoons producing rare fertile hybrids.

DNA was extracted from fin tissue samples by the Chelex/proteinase K protocol of Estoup et al. (1996). Six microsatellite markers allowing cross-priming for three Pomatoschistus species were retained (Pmic-02, Pmic-03, Pmic-07, Pmar-03, Pmar-05, Pmin-05). DThe 5' end of one of the two primers was covalently linked to fluorescein, Cy3, or Cy5 labels. Polymerase chain reactions (PCR) were performed in an Eppendorf Mastercycler programmable thermocycler with a 10 µL reaction total volume containing 0.1 U of Taq polymerase (Sigma-Aldrich), 2.5 mM MgCl₂, 0.4 mM of each dNTP (Invitrogen), 1 µL of 10X reaction buffer and 0.75 µM of each primer (Eurofins MWG). The thermal cycle was composed of an initial denaturation (94°C, 5 min); then of 34 repetitions of denaturation (94°C, 45 s), annealing (45 s at the temperatures given in Table 3 for each locus) and extension (72°C, 45 s); and then a final extension (72°C, 5 min).

Separation of amplified DNA fragments according to their size was carried out in a polyacrylamide denaturing gel 8% (Bio-Rad). The PCR products were visualized on a Hitachi FMBIO-II fluorescent imaging system (Hitachi) scanner. Allele size was determined by comparison with a fluorescently labelled ladder of known size (100–600 bp, Promega), with the help of an image analysis software FMBIO ANALYSIS 8.0 (Hitachi).

Genotypes are given with the two alleles gathered (ex: 120124). Missing data is indicated with a zero.