Metschnikowia hamakuensis sp. nov., Metschnikowia kamakouana sp. nov. and Metschnikowia mauinuiana sp. nov., three endemic yeasts from Hawaiian nitidulid beetles Marc-Andre ´ Lachance, 1 Curtis P. Ewing, 2 Jane M. Bowles 1 and William T. Starmer 3 Correspondence Marc-Andre ´ Lachance lachance@uwo.ca 1 Department of Biology, University of Western Ontario, London, Ontario, Canada N6A 5B7 2 Department of Entomology, Cornell University, Ithaca, NY 14853, USA 3 Department of Biology, Syracuse University, Syracuse, NY 13244, USA Three heterothallic, haplontic yeast species, Metschnikowia hamakuensis, Metschnikowia kamakouana and Metschnikowia mauinuiana, are described from isolates associated with endemic nitidulid beetles living on various endemic plants on three Hawaiian islands. As morphospecies, they are similar to Metschnikowia hawaiiensis, but based on mating compatibility and ascospore formation, they can be assigned clearly to distinct biological species. Analysis of ITS/5?8S and D1/D2 large subunit rDNA sequences shows that, with M. hawaiiensis and two other isolates, these species form a distinct subclade within the large-spored Metschnikowia species, indicating that they are Hawaiian endemics. Type cultures are: M. hamakuensis, UWOPS 04-207.1 T =CBS 10056 T =NRRL Y-27834 T (type, h + ) and UWOPS 04-204.1=CBS 10055=NRRL Y-27833 (allotype, h ” ); M. kamakouana, UWOPS 04-112.5 T =CBS 10058 T =NRRL Y-27836 T (type, h + ) and UWOPS 04-109.1=CBS 10057=NRRL Y-27835 (allotype, h ” ); and M. mauinuiana, UWOPS 04-190.1 T =CBS 10060 T =NRRL Y-27838 T (type, h + ) and UWOPS 04-110.4=CBS 10059=NRRL Y-27837 (allotype, h ” ). INTRODUCTION Metschnikowia and related species associated with nitidulid beetles have been discovered at an accelerating pace since the description of the first member of the large-spored group, 15 years ago (Lachance et al., 1990, 1998a, b, 2001a, 2003a). Recently, Lachance et al. (2003b) suggested that it may be possible to recognize, among Hawaiian members of the group, some that were introduced through human activity (Metschnikowia lochheadii and Candida ipomoeae) and others that may be endemic to Hawai’i (Metschnikowia hawaiiensis and Candida kipukae). In the first instance, the yeasts are associated with a Central American beetle species and adapted to warm temperatures, which would account in part for their occurrence in lowland habitats across the Hawaiian archipelago (Lachance et al., 2001a, 2003b). Show- ing conclusively that a yeast species occurs only in Hawai’i would be difficult and so the demonstration of endemism requires a different approach. For example, the observation that M. hawaiiensis is associated with a beetle that is endemic to the island of Hawai’i would support the view that the yeast itself has speciated in that region. It happens that the beetle reported as ‘nitidulid B’ by Lachance et al. (2003b) has now been identified as Prosopeus subaeneus, which is indeed a Hawaiian endemic (Ewing, 2004). Further evidence that the yeast is not an introduced exotic would arise from the isolation of a sister species of M. hawaiiensis in a similar Hawaiian habitat. In June 2004, we sampled several species of endemic nitidulids found on native plants in various nature reserves on the islands of Moloka’i, Maui and Hawai’i. Many of the isolates represented novel species, including five sister species to M. hawaiiensis. Three of these were represented by multiple strains and included both mating types; we now describe them as Metschnikowia hamakuensis sp. nov., Metschnikowia kamakouana sp. nov. and Metschnikowia mauinuiana sp. nov. METHODS Collections. The collection sites relevant to this study (Table 1) were the following Hawaiian rainforest localities: Kamakou Preserve area (21u 89 N 156u 559 W, 1200 m), Moloka’i; Waikamoi Preserve area (20 489 N 156u 159 W, 1300 m), Maui; upper Hamakua ditch (20u 59 N 155u 419 W, 1200 m), Hawai’i; Pu’u Pala and Pu’u Published online ahead of print on 14 January 2005 as DOI 10.1099/ ijs.0.63615-0. The GenBank/EMBL/DDBJ accession numbers for the ITS/5?8S and D1/D2 LSU rDNA sequences of the novel isolates identified in this study are shown in Fig. 2. 63615 G 2005 IUMS Printed in Great Britain 1369 International Journal of Systematic and Evolutionary Microbiology (2005), 55, 1369–1377 DOI 10.1099/ijs.0.63615-0