Small diatoms (Bacillariophyta) in cultures from the Styx River, New Zealand, including descriptions of three new species Author Novis, Phil M. Allan Herbarium, Landcare Research, P. O. Box 40, Lincoln 7640, New Zealand Author Braidwood, Jasmine Allan Herbarium, Landcare Research, P. O. Box 40, Lincoln 7640, New Zealand Author Kilroy, Cathy National Institute of Water and Atmospheric Research, P. O. Box 8602, Christchurch, New Zealand text Phytotaxa 2012 2012-08-24 64 1 11 45 http://biotaxa.org/Phytotaxa/article/view/phytotaxa.64.1.3 journal article 10.11646/phytotaxa.64.1.3 1179-3163 5062002 Nitzschia palea (Kütz.) Smith 1856 ( Figs 52–64 , 162 ) Valves linear-lanceolate, 2.8–4.4 µm wide, 24.2–33.3 µm long, with rostrate to subcapitate ends. Striae transverse, uniseriate, 41–45 in 10 µm , comprising round areolae. Fibulae spaced approximately evenly along the keel, 12–17 in 10 µm ; raphe continuous, polar ends hook-shaped over valve apices; central node and central raphe endings absent ( Fig. 63 ). Living cells contained 2–4 plate-like parietal chloroplasts, arranged around the frustule perimeter and separated at the cell centre ( Fig. 52 ). Reference: — Trobajo et al. 2009 , p. 449, table 3, figs 2–26. Specimens examined: —CHR618410!, CHR618411!, CHR618412! (cleaned frustules from cultures LCR-S:6:4:1, 12:1:2, and 29:2 respectively). Distribution: —The species is regarded as cosmopolitan and a useful bioindicator. However, the cryptic diversity discovered within it suggests that its growth preferences could currently be misunderstood. It appeared in cultures from 2 sites in the Styx (2 and 7). Molecular data: — Rbc L fragments were obtained for 3 strains from the Styx: strain LCR-S:6:4:1, 576 bp; strain LCR-S:12:1:2, 575 bp; and strain LCR-S:29:2, 571 bp. The first and last strains shared identical sequences; the other strain differed by 1 bp. The closest matches outside these strains were strain Spain C and the diatom-derived endosymbiont of Durinskia baltica (Levander) Carty & Cox 1986 (p-distances = 0.009). However, the Styx strains formed a robust monophyletic group in both the Bayesian and MPB analyses, separate from all other strains of N. palea in Genbank ( Fig. 162 ). The overall dataset was 1470 bp long, with 214 variable sites (103 parsimony informative, 42 occurring within the Styx fragments). The model chosen and implemented in the Bayesian analysis was T92+G. One fragment of 18S sequence was obtained, for strain LCR-S:6:4:1, 1243 bp. The closest matches to this sequence were the N. palea clones NPAL2, 3, 8, 9, and 12, and NZSP (from Luxembourg and Ukraine ; all p-distance = 0.001). The p-distance between the Styx strain and the remaining 3 N. palea clones in the database was 0.002. The Styx strain formed a robust clade with all these clones in both Bayesian and MPB analyses, but relationships within this clade were not resolved (presumably due to insufficient variation). The overall dataset was 1707 bp long, with 91 variable sites (39 parsimony informative, 28 occurring in the Styx fragment). The model chosen and implemented in the Bayesian analysis was K2+G+I. FIGURE 52–64. Nitzschia palea , strain LCR-S:6:4:1, CHR618410. Scales: Fig. 52, 10 µm; Fig. 61, 5 µm (use for Figs 53–62); Fig. 63, 0.5 µm; Fig. 64, 1 µm. Fig. 52. Live cells viewed by LM. Figs 53–62. Cleaned frustules viewed by LM, showing size variation and raphe canals on opposite margins (Figs 53, 54 are single valves; raphe is visible in Fig. 53). Fig. 63. TEM of cleaned frustule, centre of valve, showing continuous raphe with no central termini. Fig. 64 . SEM of cleaned frustule, showing fine areolae and lack of median fibulae. Observations :—The morphology of the Styx strains is closest to the strain Japan A2 of Trobajo et al. (2009) in terms of fibulae and striae per 10 µm , and closest to strain Spain A 3 in length range. Unfortunately, an rbc L sequence for Japan A2 is not available, and Spain A3 is not united with the Styx strains in the phylogenetic analysis. One possible conclusion from our molecular analyses is that the Styx strains form a monophyletic group as an artefact of their short sequence lengths; however, analyses in which the dataset is completely cropped to the length of the Styx sequences maintain the relationships. Trobajo et al. (2009 , 2010 ) propose that N. palea will need to be split into 3 or more species; problematically, this seems to require species to be defined purely according to molecular data. This would create serious problems for those required to identify these diatoms by LM or even SEM (i.e. most identifications), leading to the continued use of the name N. palea by default. This species shows the most convincing biogeographic structure of all the taxa determined in this study, thanks to its relatively extensive sampling. However, it is possible that the UK strain of Trobajo et al. (2010) would form a clade with the Styx strains, since it occupies a similar position in a cox 1 analysis ( rbc L data are not available for this strain). Order Cymbellales Family Gomphonemataceae