Some enchytraeid species of tropical, subtropical or Mediterranean origin can appear in artificial environments,e.g. green houses due to the worldwide commercial network. Since the used soil from green houses is often disposed outdoors, aquestion raised that these exotic enchytraeid species could survive under continental climate conditions. In this experiment two ofthe resettled green house species survived outdoors the winter frost and the arid summer season in Hungary

Boros, G.

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 Opusc. Zool. Budapest, 2011, 42(2): 207–210     1Gergely Boros, Systematic Zoology Research Group of  Hungarian Academy of Sciences at the Eötvös Loránd University, H-1117 Budapest, Pázmány Péter sétány 1/C, Hungary. E-mail: henlea@gmail.com  Outdoor surviving experiment with three green house enchytraeid species (Oligochaeta: Enchytraeidae)   G. BOROS1   Abstract. Some enchytraeid species of tropical, subtropical or Mediterranean origin can appear in artificial environments, e.g. green houses due to the worldwide commercial network. Since the used soil from green houses is often disposed outdoors, a question raised that these exotic enchytraeid species could survive under continental climate conditions. In this experiment two of the resettled green house species survived outdoors the winter frost and the arid summer season in Hungary. Keywords. Pot-worms, exotic species, introduction, survival.   INTRODUCTION  evelopment of human civilization entailed the development and growth of worldwide commercial network. Due to this process, several animal species were propagated accidentally which have not been able to spread under natural circumstances. The expansion of certain oligo-chaete species is an expressive example of this phenomenon. Importation of foreign earthworm or enchytraeid species is hardly perceptible. Apart from their hidden lifestyle, as saprophagous ani-mals, occasional species exchange is not nec-essarily observable in the effect produced on en-vironment, since native and introduced species can decompose similarly.  In spite of this fact, frequent monitoring de-tected invasive earthworm species in a number of cases: Csuzdi and Szlávecz (2003) have recently recorded the exotic species Lumbricus friendi Cognetti, 1904 in North America, previously known only from the Atlantic region of Europe (Bouché, 1972). Dichogaster bolaui (Michaelsen, 1891), which is of Eastern African origin, was found not only in green houses but also in bath-rooms in Hungary and Israel (Csuzdi et al,. 2008). The same species have repeatedly been found in the sewage systems of buildings in Sweden and Finland as well (Terhivuo, 1990). Such species were defined as a ‘domicole’ which are able (or adapted) to live in urban dwellings. Earlier, Mi-chaelsen (1903), Lee (1987) and others used the expression ‘peregrine’ for earthworm species which are anthropochorous and are usually con-ﬁned to disturbed, human-modiﬁed habitats, such as gardens, lawns or green houses. The green-house invertebrate fauna of Hungary have pre-viously been investigated by Korsós et al. (2002) reporting on Isopods and Diplopods and Hídvégi (1994) who examined earthworms. Both inves-tigations demonstrated the presence of introduced peregrine species.  Despite of the broad range of the aforemen-tioned literature, enchytraeids have not yet been the subject of similar studies, albeit their presence is more evident in consequence of their size. In 2006 a comprehensive study was launched to investigate the Enchytraeid fauna of green houses in Hungary. The aim was to describe the enchyt-raeid fauna in a sample of 10 green houses across Hungary and to prove the presence of exotic species. The hypothesis of the research was that (sub)tropical and Mediterranean enchytraeid spe-cies were imported in earth-balls left on the roots of the plants, or even more among the roots them-selves, and the species settled down due to the human-made optimal conditions found in green houses. As a result, some species detected earlier in subtropical or Mediterranean climate were found in the samples, e.g. Fridericia pretoriana Stephenson, 1930, Enchytraeus dudichi Dózsa-Farkas, 1995, moreover, 2 unidentified species D  Boros: Outdoor surviving experiment with enchytraeids     208 belonging to the genus Hemienchytraeus were described (Boros and Dózsa-Farkas, 2007). A new species to science, Marionina scintillans Boros and Dózsa-Farkas, 2008 was also discovered (Bo-ros and Dózsa-Farkas, 2008). The abundance of this species was extremely high: ca. 342.000 ind./m2. This high number can be explained with the artificial circumstances, since the highest ab-undance of enchytraeids in the nature was 289.000 ind./m2 (Peachey, 1963). Since the used soil from green houses is often disposed outdoors, the question raised by this paper is whether in-troduced exotic enchytraeid species could survive under continental climate conditions, especially with that high abundance. In the course of outdoor sampling in botanical gardens, has not been found an evidence of surviving exotic populations, therefore an experiment was carried out to answer this question.  This experiment was carried out in the Bo-tanical Garden of the Eötvös Loránd University. Three species were chosen: Enchytraeus bigemi-nus Nielsen & Christensen, 1963; the above men-tioned  Marionina scintillans, and a fragmenting Marionina species found in the botanical garden of Szeged University in 2009 and uniquely in the genus able to multiply by architomy. Although stable breeding cultures are sustained, no sexually matured specimens of this species are known yet. Even so, the species was considered as suited to this study due to the special asexual reproduction like that of E. bigeminus.  MATERIAL AND METHODS  The research were carried out in the Botanical Garden of the Eötvös Loránd University. A 0.25 m2 (0.5x0.5 m) undisturbed lawn area was en-closed where lack of any gardening intervention was guaranteed. Before resettlement, qualitative samples were taken from this area to establish the present Enchytraeid species.  Individuals of the 3 species were placed in the following numbers: 100 specimens each of  En-chytraeus bigeminus and Marionina scintillans, and 40 specimens of the fragmenting Marionina   species. Breeding cultures of the fragmenting Ma-rionina sp. increase slowly, so only a small num-ber of worms were to be expended.   The reasons why these species were chosen are 1) these species were missing from the research area so the presence of the newly introduced species were traceable 2) due to reliable breeding cultures relatively high number of enchytraeids were available.   Breeding cultures were sustained in Petri-dishes on 1% agar-agar medium with soil on top, earlier sterilized by heat-treatment. Foods were rolled oats and dried leaves of nettle (Urtica dioica). Breeding cultures were kept between 10–12 °C in refrigerator.)  Quantitative samples were taken 5 times in the year of 2011: 2 times both in spring (09. 04. and 28. 04.) and in summer (24. 06. and 11. 07.) and once in autumn (06. 09.). Every time 5 samples were taken with a 20 cm2 surface split soil corer. Samples were 10 cm deep which means 200 cm3 volumes each.  Enchytraeids were extracted from the soil by the wet funnel method (O’Connor, 1962) and were identified alive with a Zeiss Axioskop Imager.A2 microscope, using DIC (differential interference contrast) illumination. Voucher spe-cimens were preserved in 70% ethanol and stored in 600 µl. Eppendorf tubes.  RESULTS  Just before the resettlement (05. 04. 2010.) six species of five genera were found in the research field:  Achaeta eiseni Vejdovský, 1878 Buchholzia appendiculata (Buchholz, 1862) Enchytraeus buchholzi Vejdovský, 1879 Enchytraeus sp. juv. Fridericia bulboides Nielsen & Christensen, 1959 Henlea ventriculosa (d’Udekem, 1854)   Boros: Outdoor surviving experiment with enchytraeids     209It was impossible to identify specimens of En-chytraeus sp. juv. due to the lack of sexual organs. However, based on setal complements and the number of setal follicles, it was obvious that this species is identical neither with E. bigeminus nor E. buchholzi.  The following species were found in the spring samples (recollected species are represented in bold):  Buchholzia appendiculata Enchytraeus bigeminus Enchytraeus buchholzi Enchytraeus sp. juv. Fridericia bulboides Henlea ventriculosa Marionina scintillans  Two of the resettled species were present which justify their survival of the winter frost. All specimens of Marionina scintillans were adults, in some of them matured egg was observable. At the same time, individuals of the other Marionina species completely disappeared.  Species found during the summer sampling were:  Enchytraeus bigeminus Fridericia bulboides Enchytraeus buchholzi  Low numbers of individuals were present from all the three species. In some samples there were absolutely no enchytraeids.   Species found during the autumn sampling were:  Buchholzia appendiculata Enchytraeus bigeminus Enchytraeus buchholzi Fridericia bulboides Marionina scintillans  Moisture demanding Buchholzia appendicula-ta and M. scintillans reappeared in the samples, but only in low numbers.  From the original fauna of the research field, Achaeta eiseni were not found in any samples after introduction of the 3 new species.  DISCUSSION  The number of species of the research area was relatively low compared to other lawns in the region, e.g. 5–15 species in the meadows near Tisza river, Hungary (Dózsa-Farkas et al., 2003), 7–12 species in lawn parks of Brno, Czech Re-public (Schlaghamerský and Pižl, 2009) or 6–10 species in the grasslands of Sas-hegy Nature Conservation Area, Budapest, Hungary (Boros, 2007). This low number of species was especially prominent in the dry summer season when spe-cimen numbers were also small. The periodical lack of Buchholzia appendiculata also confirms the importance of aridity in the seasonal deve-lopment of species’ abundances which is also in line with earlier findings.  In the course of the experiment two displaced species were found outdoors. E. bigeminus is well known from compost fields, and is considered as a common species in Southern Europe (Schmelz and Collado, 2010). Now, it seems we can find it in colder continental climate as well. Expressly, an advantage is that this species can multiply asexually by fragmentation (architomy) so in-creases their populations quickly. Earlier expe-riences showed that this species can contaminate other enchytraeid breeding cultures so it can be an invasive species.  M. scintillans was so far known only from green houses. However, this experiment proved its ability to survive outdoors as well. This species was missing from the samples in the summer but reappeared during the moist autumn. A presu-mable explanation for this behaviour is that only cocoons survived the arid period and all the worms died. Another possibility is that these worms go deeper than the sampling depth of 10 cm during the arid season.  The fragmenting Marionina species was not found outdoors in the course of the study which possibly means the extinction of these enchyt-raeids during the harsh winter conditions. Another  Boros: Outdoor surviving experiment with enchytraeids     210 explanation could be the lower initial number of the test worms. As it was already mentioned, Achaeta eiseni was also missing in every sample, in spite of the fact it was present in the study area before. This phenomenon should be placed under more thor-ough scrutiny in the future.  The preliminary results of the experiment herewith presented show that several green house species are able to survive outdoors under con-tinental climate. For the more reliable explanation of their behaviour and survival further expe-riments are needed with higher number of en-chytraeid species and individuals.  Acknowledgements – The author is grateful to Dr. László Orlóczy, Director of the ELTE Botanical Garden for providing an opportunity to execute this study. Many thanks are due to Márton Czirfusz (HAS Centre for Regional Studies) for correcting the English of this paper. 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Outdoor surviving experiment with three green house enchytraeid species (Oligochaeta: Enchytraeidae)

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