Sofie Derycke

Dispersal and gene flow in free-living marine nematodes

Dispersal and gene flow determine connectivity among populations, and can be studied through population genetics and phylogeography. We here review the results of such a framework for free-living marine nematodes. Although field experiments have illustrated substantial dispersal in nematodes at ecological time scales, analysis of the genetic diversity illustrated the importance of priority effects, founder effects and genetic bottlenecks for population structuring between patches <1 km apart. In contrast, only little genetic structuring was observed within an estuary (<50 km), indicating that these small scale fluctuations in genetic differentiation are stabilized over deeper time scales through extensive gene flow. Interestingly, nematode species with contrasting life histories (extreme colonizers vs persisters) or with different habitat preferences (algae vs sediment) show similar, low genetic structuring. Finally, historical events have shaped the genetic pattern of marine nematodes and show that gene flow is restricted at large geographical scales. We also discuss the presence of substantial cryptic diversity in marine nematodes, and end with highlighting future important steps to further unravel nematode evolution and diversity.

Phylogeography of the **Rhabditis (Pellioditis) marina** species complex: evidence for long-distance dispersal, and for range expansions and restricted gene flow in the northeast Atlantic

Pinpointing processes that structure the geographical distribution of genetic diversity of marine species and lead to speciation is challenging because of the lack of obvious dispersal barriers and the likelihood of substantial (passive) dispersal in oceans. In addition, cryptic radiations with sympatric distributions abound in marine species, challenging the allopatric speciation mechanism. Here, we present a phylogeographical study of the marine nematode species complex Rhabditis (Pellioditis) marina to investigate processes shaping genetic structure and speciation. Rhabditis (P.) marina lives on decaying macroalgae in the intertidal, and may therefore disperse over considerable distances. Rhabditis (P.) marina consists of several cryptic species sympatrically distributed at a local scale. Genetic variation in the COI gene was screened in 1362 specimens from 45 locations around the world. Two nuclear DNA genes (ITS and D2D3) were sequenced to infer phylogenetic species. We found evidence for ten sympatrically distributed cryptic species, seven of which show a strong genetic structuring. A historical signature showed evidence for restricted gene flow with occasional long‐distance dispersal and range expansions pre‐dating the last glacial maximum. Our data also point to a genetic break around the British Isles and a contact zone in the Southern Bight of the North Sea. We provide evidence for the transoceanic distribution of at least one cryptic species (PmIII) and discuss the dispersal capacity of marine nematodes. The allopatric distribution of some intraspecific phylogroups and of closely related cryptic species points to the potential for allopatric speciation in R. (P.) marina.

Exploring the Use of Cytochrome Oxidase c Subunit 1 (COI) for DNA Barcoding of Free-Living Marine Nematodes

Background The identification of free-living marine nematodes is difficult because of the paucity of easily scorable diagnostic morphological characters. Consequently, molecular identification tools could solve this problem. Unfortunately, hitherto most of these tools relied on 18S rDNA and 28S rDNA sequences, which often lack sufficient resolution at the species level. In contrast, only a few mitochondrial COI data are available for free-living marine nematodes. Therefore, we investigate the amplification and sequencing success of two partitions of the COI gene, the M1-M6 barcoding region and the I3-M11 partition. Methodology Both partitions were analysed in 41 nematode species from a wide phylogenetic range. The taxon specific primers for the I3-M11 partition outperformed the universal M1-M6 primers in terms of amplification success (87.8% vs. 65.8%, respectively) and produced a higher number of bidirectional COI sequences (65.8% vs 39.0%, respectively). A threshold value of 5% K2P genetic divergence marked a clear DNA barcoding gap separating intra- and interspecific distances: 99.3% of all interspecific comparisons were >0.05, while 99.5% of all intraspecific comparisons were <0.05 K2P distance. Conclusion The I3-M11 partition reliably identifies a wide range of marine nematodes, and our data show the need for a strict scrutiny of the obtained sequences, since contamination, nuclear pseudogenes and endosymbionts may confuse nematode species identification by COI sequences.

Linking DNA sequences to morphology: cryptic diversity and population genetic structure in the marine nematode Thoracostoma trachygaster (Nematoda, Leptosomatidae)

Derycke, S., De Ley, P., De Ley, I.T., Holovachov, O., Rigaux, A. & Moens, T. (2010). Linking DNA sequences to morphology: cryptic diversity and population genetic structure in the marine nematode Thoracostoma trachygaster (Nematoda, Leptosomatidae).—Zoologica Scripta, 39, 276–289.

Influence of bacterivorous nematodes on the decomposition of cordgrass

The influence of bacterivorous nematodes (Diplolaimelloides meyli, Diplolaimelloides oschei, Diplolaimella dievengatensis, Panagrolaimus paetzoldi) on the decomposition of a macrophyte (Spartina anglica) in an aquatic environment was investigated by using laboratory microcosm experiments. Several earlier studies have shown enhancement of the decomposition process in the presence of nematodes, but nematode species-specific effects were never tested. In this study four bacterivorous nematode species were applied separately to microcosms to investigate such speciesspecific influences. No stimulation of the decomposition process nor of the microbial community was observed in the presence of the nematodes, both were highest in the absence of nematodes. However, clear differences were found between nematode treatments. P. paetzoldi reached much higher numbers than the other species, causing a decrease in microbial activity, probably due to (over)grazing. Remarkably this low microbial activity did not result in a slow-down of the decomposition process compared to the other nematode treatments, raising the question whether P. paetzoldi might be able to directly assimilate detrital compounds. Other nematode species reached much lower densities, but nevertheless an influence on the decomposition process was observed. However, this experiment does not support the view that bacterivorous nematodes enhance decomposition rate. The experimental results show that in nematode communities the use of functional groups is inadequate for biodiversity studies. The four nematode species used in this study belong to the same functional group, but are clearly not functionally redundant since they all have a different influence

Species boundaries and phylogenetic relationships between Atlanto-Mediterranean shallow-water and deep-sea coral associated Hexadella species (Porifera, Ianthellidae)

Coral reefs constitute the most diverse ecosystem of the marine realm and an increasing number of studies are focusing on coral species boundaries, distribution, and on processes that control species ranges. However, less attention has been paid to coral associated species. Deep-sea sponges dominate cold-water coral ecosystems, but virtually nothing is known about their molecular diversity. Moreover, species boundaries based on morphology may sometimes be inadequate, since sponges have few diagnostic characters. In this study, we investigated the molecular diversity within the genus Hexadella (Porifera, Demospongiae, Verongida, Ianthellidae) from the European shallow-water environment to the deep-sea coral ecosystems. Three molecular markers were used: one mitochondrial (COI) and two nuclear gene fragments (28S rDNA and the ATPS intron). Phylogenetic analyses revealed deeply divergent deep-sea clades congruent across the mitochondrial and nuclear markers. One clade contained specimens from the Irish, the Scottish, and the Norwegian margins and the Greenland Sea (Hexadella dedritifera) while another clade contained specimens from the Ionian Sea, the Bay of Biscay, and the Irish margin (H. cf. dedritifera). Moreover, these deeply divergent deep-sea clades showed a wide distribution suggesting a connection between the reefs. The results also point to the existence of a new deep-sea species (Hexadella sp.) in the Mediterranean Sea and of a cryptic shallow-water species (Hexadella cf. pruvoti) in the Gorringe Bank. In contrast, low genetic differentiation between H. cf. dedritifera and H. pruvoti from the Mediterranean Sea was observed. All Hexadella racovitzai specimens from the Mediterranean Sea (shallow and deep) to the Atlantic formed a monophyletic group.

Coexisting cryptic species of the Litoditis marina complex (Nematoda) show differential resource use and have distinct microbiomes with high intraspecific variability

Differences in resource use or in tolerances to abiotic conditions are often invoked as potential mechanisms underlying the sympatric distribution of cryptic species. Additionally, the microbiome can provide physiological adaptations of the host to environmental conditions. We determined the intra‐ and interspecific variability of the microbiomes of three cryptic nematode species of the Litoditis marina species complex that co‐occur, but show differences in abiotic tolerances. Roche 454 pyrosequencing of the microbial 16S rRNA gene revealed distinct bacterial communities characterized by a substantial diversity (85–513 OTUs) and many rare OTUs. The core microbiome of each species contained only very few OTUs (2–6), and four OTUs were identified as potentially generating tolerance to abiotic conditions. A controlled experiment in which nematodes from two cryptic species (Pm1 and Pm3) were fed with either an E. coli suspension or a bacterial mix was performed, and the 16S rRNA gene was sequenced using the MiSeq technology. OTU richness was 10‐fold higher compared to the 454 data set and ranged between 1118 and 7864. This experiment confirmed the existence of species‐specific microbiomes, a core microbiome with few OTUs, and high interindividual variability. The offered food source affected the bacterial community and illustrated different feeding behaviour between the cryptic species, with Pm3 exhibiting a higher degree of selective feeding than Pm1. Morphologically similar species belonging to the same feeding guild (bacterivores) can thus have substantial differences in their associated microbiomes and feeding strategy, which in turn may have important ramifications for biodiversity–ecosystem functioning relationships.

The link between microbial diversity and nitrogen cycling in marine sediments is modulated by macrofaunal bioturbation

Objectives The marine benthic nitrogen cycle is affected by both the presence and activity of macrofauna and the diversity of N-cycling microbes. However, integrated research simultaneously investigating macrofauna, microbes and N-cycling is lacking. We investigated spatio-temporal patterns in microbial community composition and diversity, macrofaunal abundance and their sediment reworking activity, and N-cycling in seven subtidal stations in the Southern North Sea. Spatio-Temporal Patterns of the Microbial Communities Our results indicated that bacteria (total and β-AOB) showed more spatio-temporal variation than archaea (total and AOA) as sedimentation of organic matter and the subsequent changes in the environment had a stronger impact on their community composition and diversity indices in our study area. However, spatio-temporal patterns of total bacterial and β-AOB communities were different and related to the availability of ammonium for the autotrophic β-AOB. Highest bacterial richness and diversity were observed in June at the timing of the phytoplankton bloom deposition, while richness of β-AOB as well as AOA peaked in September. Total archaeal community showed no temporal variation in diversity indices. Macrofauna, Microbes and the Benthic N-Cycle Distance based linear models revealed that, independent from the effect of grain size and the quality and quantity of sediment organic matter, nitrification and N-mineralization were affected by respectively the diversity of metabolically active β-AOB and AOA, and the total bacteria, near the sediment-water interface. Separate models demonstrated a significant and independent effect of macrofaunal activities on community composition and richness of total bacteria, and diversity indices of metabolically active AOA. Diversity of β-AOB was significantly affected by macrofaunal abundance. Our results support the link between microbial biodiversity and ecosystem functioning in marine sediments, and provided broad correlative support for the hypothesis that this relationship is modulated by macrofaunal activity. We hypothesized that the latter effect can be explained by their bioturbating and bio-irrigating activities, increasing the spatial complexity of the biogeochemical environment.

Phylogenetic Relationships among NE Atlantic Plocamionida Topsent (1927) (Porifera, Poecilosclerida): Under-Estimated Diversity in Reef Ecosystems

Background Small and cryptic sponges associated with cold-water coral reefs are particularly numerous and challenging to identify, but their ecological and biochemical importance is likely to compete with megabenthic specimens. Methodology/Principal Findings Here we use a combination of the standard M1M6 and I3M11 partitions of the COI fragment, partial rDNA 28S sequences and morphology to delineate small encrusting Plocamionida species. In total, 46 specimens were retrieved from seven shallow to deep-water coral locations, crossing 3,000 km along the European margins. Our work provides evidence that the Plocamionida ambigua f. tylotata and f. grandichelata can be considered valid species, whereas Plocamionida ambigua f. tornata corresponds to the species P. ambigua. Within the monophyletic group of Plocamionida, P. microcionides is shown as really divergent from the other taxa, and four putative new Plocamionida species are suggested. Conclusions/Significance This study shows that the use of molecular and morphological information in an integrative approach is a powerful tool for the identification of sponge species, and suggests that an under-estimated biodiversity of sponges occurs in cold-water coral reefs.

Systematics and DNA barcoding of free-living marine nematodes with emphasis on tropical desmodorids using nuclear SSU rDNA and mitochondrial COI sequences

The diversity and phylogenetic relationships of the Desmodoridae, a widespread tropical family of free-living marine nematodes, is hitherto poorly known both from molecular and taxonomic points of view. We performed a molecular phylogenetic analysis of marine nematodes to: i) disentangle relationships among tropical desmodorid species; and ii) compare the performance of the nuclear SSU rDNA and mitochondrial COI nucleotide sequences in 42 and 45 nominal species, respectively, to identify species. We generated 27 new sequences of SSU rDNA belonging to five genera not previously sequenced, and 34 new sequences of COI belonging to six genera and four families not previously sequenced. The SSU rDNA tree confirmed the Enoplida to be a monophyletic sister group to the Chromadorida. The family Comesomatidae is a sister group of the Xyalidae within the Monhysterida. Both DNA markers confirmed the congruence between the morphology- and molecular-based phylogenetic inferences for most of the families. Desmodoridae was a monophyletic group, but the relationships within the family could not be recovered; the subfamilies Desmodorinae and Spiriniinae were not monophyletic meanwhile the monophyly of Stilbonematinae was not fully supported due to a few specimens of questionable identity. COI performed better than SSU rDNA to disentangle relationships among closely related species and suggested the presence of cryptic diversity within Desmodoridae. COI is effective to explore cryptic diversity and barcode species within Nematoda, with a possible threshold of genetic distance of 5% between conspecific and interspecific sequences, but DNA barcoding is limited by the poor knowledge of the diversity and taxonomy of the group and the lack of a good reference database of vouchered COI sequences.