Molecular identification key based on PCR/RFLP for three polychaete sibling species of the genus Marenzelleria, and the species’ current distribution in the Baltic Sea

Abstract

Studies of Marenzelleria species were often hampered by identification uncertainties when using morphological characters only. A newly developed PCR/RFLP protocol allows a more efficient discrimination of the three species Marenzelleria viridis, Marenzelleria neglecta and Marenzelleria arctia currently known for the Baltic Sea. The protocol is based on PCR amplification of two mitochondrial DNA gene segments (16S, COI) followed by digestion with restriction enzymes. As it is faster and cheaper than PCR/sequencing protocols used so far, the protocol is recommended for large-scale analyses. The markers allow an undoubted determination of species irrespective of life stage or condition of the worms in the samples. The protocol was validated on about 950 specimens sampled at more than 30 sites of the Baltic and the North Sea, and on specimens from populations of the North American east coast. Besides this test we used mitochondrial DNA sequences (16S, COI, Cytb) and starch gel electrophoresis to further investigate the distribution of the three Marenzelleria species in the Baltic Sea. The results show that M. viridis (formerly genetic type I or M. cf. wireni) occurred in the Ö resund area, in the south western as well as in the eastern Baltic Sea, where it is found sympatric with M. neglecta. Allozyme electrophoresis indicated an introduction by range expansion from the North Sea. The second species, M. arctia, was only found in the northern Baltic Sea, where it sometimes occurred sympatric with M. neglecta or M. viridis. For Baltic M. arctia, the most probable way of introduction is by ship ballast water from the European Arctic. There is an urgent need for a new genetic analysis of all Marenzelleria populations of the Baltic Sea to unravel the current distribution of the three species.

Are similar worms different? A comparative tracer study on bioturbation in the three sibling species Marenzelleria arctia, M. viridis, and M. neglecta from the Baltic Sea

“…Additional layers were taken at 16 and 20 cm (C); at 12, 14, 16, 18, and 20 cm (MA); and at 35 cm (MN and MV). The residence depth of polychaetes was noted during the sectioning, and collected worms were preserved in 80% ethanol for identification according to Blank and Bastrop (2009). Sediment layers were weighed, and pore water was extracted by centrifuging for 1 min at 500 rpm in cups with GF/F filters as described by Saager et al (1990).…”

“…The spionid polychaetes of the genus Marenzelleria consist of six species (Fauchald and Bellan 2012) with the three sibling species M. neglecta, M. viridis, and M. arctia present in the Baltic Sea since the 1980s (Bick and Burckhardt 1989;Bastrop 1997;Blank et al 2008). Sibling species are two or more related species that are morphologically difficult or impossible to distinguish (Mayr and Ashlock 1991).…”

“…Considerable numbers of investigations with Marenzelleria spp. have been done so far, but Blank et al (2008) found incorrect species nominations in astonishingly 58% of all considered papers as a result of the difficulties in species discrimination. Due to the similarity in morphological characteristics, the sibling species are thought to have only minor, if any, functional differences (Hietanen et al 2007;Norkko et al 2011).…”

“…Knowledge on the distribution of the three sibling species in the Baltic assumes that M. neglecta is established over the whole Baltic Sea, whereas spreading of M. viridis and M. arctia is still an ongoing process (Blank et al 2008). M. viridis and M. neglecta originate from North America (Leppä koski and Olenin 2000) in contrast to M. arctia, which is restricted originally to the European Arctic (Jørgensen et al 1999).…”

“…M. viridis and M. neglecta originate from North America (Leppä koski and Olenin 2000) in contrast to M. arctia, which is restricted originally to the European Arctic (Jørgensen et al 1999). Range expansion from the North Sea in case of M. viridis and displacement via ship ballast water for all of the three sibling species are the most likely reasons for facilitated invasion to the Baltic Sea (Blank et al 2008).…”