Downloaded from www.microbiologyresearch.org by IP: 54.205.18.105 On: Fri, 24 Jun 2016 04:00:13 Herpetomonas trimorpha sp. nov. (Trypanosomatidae, Kinetoplastida), a parasite of the biting midge Culicoides truncorum (Ceratopogonidae, Diptera) Lenka Zı ´dkova ´, 1 Ivan Cepicka, 2 Jan Voty ´pka 1 and Milena Svobodova ´ 1 Correspondence Lenka Zı ´dkova ´ murfar@seznam.cz 1 Department of Parasitology, Faculty of Science, Charles University in Prague, Vinicna 7, Prague 128 44, Czech Republic 2 Department of Zoology, Faculty of Science, Charles University in Prague, Vinicna 7, Prague 128 44, Czech Republic Monoxenous trypanosomatid Herpetomonas trimorpha sp. nov. was isolated from the digestive tract of the biting midge Culicoides truncorum (Ceratopogonidae, Diptera). This species forms three distinct morphotypes in culture: the microflagellate promastigote, the small promastigote and the long promastigote. The last form is unique for the newly described species. Phylogenetic analyses of SSU rRNA and glycosomal glyceraldehyde phosphate dehydrogenase genes showed that H. trimorpha sp. nov. is the closest relative of Herpetomonas ztiplika, another monoxenous trypanosomatid isolated from biting midges. However, morphological and randomly amplified polymorphic DNA analyses confirmed that H. trimorpha sp. nov. is distinct from H. ztiplika. INTRODUCTION The genus Herpetomonas comprises monoxenous trypano- somatids, which mainly parasitize the digestive tract of muscid Diptera (Wallace, 1966; Molyneux, 1977; McGhee & Cosgrove, 1980; Podlipaev, 1990). However, herpeto- monads have also been isolated from plants and even mammals (Morsy et al., 1988; Catarino et al., 2001; Fiorini et al., 2001; Podlipaev et al.,2004a; Marı´n et al., 2007). The classification of the genus Herpetomonas is problematic and, similar to most other monoxenous trypanosomatid genera, the genus is polyphyletic (Camargo et al., 1992; Teixeira et al., 1997; Hollar et al., 1998; Hughes & Piontkivska, 2003; Podlipaev et al., 2004a). Therefore, a major taxonomic revision of ‘lower’ trypanosomatids is needed (see Bulat et al., 1999; Podlipaev, 2000, 2001; Maslov et al., 2001; Merzlyak et al., 2001; Momen, 2001; Yurchenko et al., 2008, 2009). Over 30 herpetomonad species have been described at the time of writing (McGhee & Cosgrove, 1980; Podlipaev, 1990). Various authors have used different approaches for the description of new species (Nunes et al., 1994; Teixeira et al., 1997; Maslov et al., 2001; Santos et al., 2005). Morphology-based taxonomy has been criticized by some authors, who argued that most trypanosomatids are morphologically too variable to possess reliable diagnostic characters (Wallace et al., 1983; Batistoti et al., 2001; Merzlyak et al., 2001; Podlipaev, 2001; Yurchenko et al., 2008, 2009). Indeed, the classical diagnostic character, i.e. the presence of promastigotes and opisthomastigotes, is no longer considered to be sufficient for assigning a trypano- somatid to the genus Herpetomonas (Camargo et al., 1992; Podlipaev et al., 2004b). On the other hand, some trypanosomatids were described solely on the basis of their host specificity, following the ‘one host – one parasite’ paradigm. However, this approach was not found univer- sally applicable as single trypanosomatid species have been shown to infect more host species and vice versa (Bulat et al., 1999; Podlipaev, 2003; Podlipaev et al., 2004a; Svobodova ´ et al., 2007). Recently, the most promising approach for species description seems to be a combination of several methods including molecular phylogenetics (Batistoti et al., 2001; Yurchenko et al., 2006; Svobodova ´ et al., 2007). Here, based on morphology, ultrastructure, phylogenetic analysis of SSU rRNA and glycosomal glyceraldehyde phosphate dehydrogenase (gGAPDH) genes, and randomly amplified polymorphic DNA (RAPD) analysis, we describe Herpetomonas trimorpha sp. nov. as a parasite of the intestinal tract of biting midges (Ceratopogonidae), closely related to Herpetomonas ztiplika. The main feature of the novel species is the ability to form three distinct morphotypes in culture. Abbreviations: gGAPDH, glycosomal glyceraldehyde phosphate dehy- drogenase; RAPD, random amplified polymorphic DNA. The GenBank/EMBL/DDBJ accession numbers for the SSU rRNA and gGAPDH gene sequences of H. trimorpha sp. nov. are EU179326 and FJ850241, respectively. The accession number for the gGAPDH gene sequence of H. ztiplika is FJ850242. Random amplified polymorphic DNA amplification patterns of isolate CER 9 and related isolates are available with the online version of this paper. International Journal of Systematic and Evolutionary Microbiology (2010), 60, 2236–2246 DOI 10.1099/ijs.0.014555-0 2236 014555 G 2010 IUMS Printed in Great Britain