Genome Evolution in the Nematode Pristionchus paciVcus

Pristionchus spp. nematodes exhibit several traits that might serve as pre-adaptations to parasitism. Under harsh environmental conditions, these nematodes can arrest development and form dauer larvae. In addition, they have been shown to live in necromenic association with a range of beetles, including dung beetles (Geotrupes stercorosus) on which, for example, Pristionchus entomophagus is commonly found. It has been argued that the formation of dauer larvae and the association with invertebrates represent intermediate steps towards parasitism. To better understand necromenic associations, and to gain information on Pristionchus spp. abundance and the general species composition on dung beetles, we extracted all the nematode fauna present on 114 individuals of G. stercorosus. By direct sequencing using the 18S SSU, we provide a barcode for all nematodes isolated from the beetle samples. In total, 5,002 dauer-stage nematodes were sequenced, which included Pristionchus spp. , Koerneria spp. (Diplogastridae), Pelodera spp. (Rhabditidae), and Strongyloidea as well as Spirurida. Intensities of infection varied from over 1,000 nematodes isolated from a single G. stercorosus to none, with Pelodera spp. being the most abundant group isolated. This study presents the first quantitative data on the Pristionchus spp. infection of beetles. Studies of nematodes from the diplogasterid genus Pristionchus have revealed both a wealth of genetic and genomic techniques and a good understanding of ecology. Physical and genetic linkage maps and a fully sequenced genome, as well as transgenic techniques, are available (Dieterich et al., 2008; Schlager et al., 2009). There are more than 150 P. pacificus isolates collected from around the world and 25 Pristionchus species are in culture. With these attributes, Pristionchus pacificus serves as a model system in evolutionary developmental biology (evo-devo) and for comparison with Caenorhabditis elegans (Zheng et al., 2005; Tian et al., 2008). More recently, P. pacificus has been used for evolutionary ecology and population genetic studies (Herrmann et al., 2007; Zauner et al., 2007). In the natural environment, most Pristionchus species are found on beetles in a species-specific manner. For example, Pristionchus maupasi and Pristionchus entomophagus are found on cockchafers (Melolontha sp.) and dung beetles (Geotrupes sp.), respectively (Herrmann et al., 2006a), while P. pacificus and Pristionchus uniformis are predominantly found on the oriental beetle (Exomala orientalis) and the Colorado potato beetle (Leptinotarsa decemlineata), respectively (Herrmann et al., 2006a, 2007). These nematodes are thought to have a necromenic relationship with their hosts (Kiontke and Sudhaus, 2006), whereby they infest the beetle and, upon beetle death, they feed on a variety of bacteria, fungi, and other nematodes that proliferate on the carcass (Rae et al., 2008). In addition, Pristionchus species display unique chemoattraction profiles when exposed to insect and plant semiochemicals (Hong and Sommer, 2006; Herrmann et al., 2007; Hong et al., 2008). These studies also add to our understanding of the evolution of parasitism, as Pristionchus spp. display several traits that may function as pre-adaptations to a parasitic lifestyle, e.g., the formation of dauer larvae, high toxicity tolerance, and low oxygen tolerance (Dieterich and Sommer, 2009). Pre-adaptations are defined as adaptations of an organism to its current environment that might be co-opted for new functions in the process of acquiring a different niche during the course of evolution (Osche, 1956). The importance of pre-adaptations in the evolution of parasitism has been discussed for many years by multiple authors, largely on hypothetical grounds (Osche 1956; Anderson, 1984; Poulin, 2007). Still, there are controversies on the term pre-adaptation and in other scientific disciplines, such as evolutionary psychology, the word ‘‘coopted adaptation’’ has been proposed instead of ‘‘pre-adaptation’’ (Buss et al., 1998). As experimental data have remained scarce, Pristionchus spp. nematodes could emerge as a model system, in this context, due to their combination of genetic tools and a necromenic host association (Dieterich and Sommer, 2009; Ogawa et al., 2009). The development of molecular tags, e.g., sequencing of 18S small subunit ribosomal RNA gene (SSU), provides a means for rapid analysis of the molecular operational taxonomic units (MOTU) present in nematode communities (Blaxter et al., 1998; Floyd et al., 2002; Griffiths et al., 2006). Using a similar molecular approach, we profiled the natural, resident nematode community present on dung beetles Geotrupes (Anoplotrupes) stercorosus L. from the Schönbuch Forest in Tübingen, Germany. These beetles are highly abundant in the forest habitat and are hosts for a large number of nematode species including Pristionchus spp. , Koerneria spp. , and Pelodera spp. (Kühne, 1996; Herrmann et al., 2006a). The dung beetle family has 3 subfamilies (Aphodiinae, Geotrupinae, and Scarabaeinae) (Janssens, 1960) and can be grouped into dwellers, tunnellers, or rollers (Cambefort and Hanski, 1991) according to dung life-style. The Geotrupinae are tunnellers and dig a vertical tunnel beneath the dung heap, then move dung to the shaft base (D’hondt et al., 2008). Larvae can remain in pupal cells for an average of 5–6 mo for G. spiniger and 9–10 mo for G. vernalis (Kühne, 1996), giving ample time for nematode colonization. The main aim of the present study was to gather information on the number and life stage of Pristionchus spp. individuals on G. stercorosus to provide a qualitative assessment of the necromenic beetle association. Additionally, we wanted to identify and quantify the rest of the nematode community. Received 2 September 2009; revised 11 November 2009, 7 December 2009; accepted 21 January 2010. *To whom correspondence should be addressed. DOI: 10.1645/GE-2319.1 J. Parasitol. , 96(3), 2010, pp. 525–531 F American Society of Parasitologists 2010