A Best Practice Example from Diatom Research

DNA barcoding uses a short fragment of a DNA sequence to identify a taxon. After obtaining the target sequence it is compared to reference sequences stored in a database to assign an organism name to it. The quality of data in the reference database is the key to the success of the analysis. In the here presented study, multiple types of data have been combined and critically examined in order to create best practice guidelines for taxonomic reference libraries for environmental barcoding. 70 unialgal diatom strains from Berlin waters have been established and cultured to obtain morphological and molecular data. The strains were sequenced for 18S V4 rDNA (the pre-Barcode for protists) as well as rbcL data, and identified by microscopy. LM and for some strains also SEM pictures were taken and physical vouchers deposited at the BGBM. 37 freshwater taxa from 15 naviculoid diatom genera were identified. Four taxa from the genera Amphora, Mayamaea, Planothidium and Stauroneis are described here as new. Names, molecular, morphological and habitat data as well as additional images of living cells are also available electronically in the AlgaTerra Information System. All reference sequences (or reference barcodes) presented here are linked to voucher specimens in order to provide a complete chain of evidence back to the formal taxonomic literature

Integrative taxonomic description of two new species of the Cocconeis placentula group (Bacillariophyceae) from Korea based on unialgal strains

Cocconeis coreana and C. sijunghoensis are described as new based on micromorphological and molecular data C. coreana is represented by five unialgal cultures from four different freshwater bodies, two from North Korea and three from South Korea. C. sijunghoensis is represented by two unialgal cultures from a brackish water body in North Korea. Except for one, all of the strains auxosporulated and showed an almost quadrupling of size in length and width. Morphologically, these species with their two different elliptical valves belong to the Cocconeis placentula group. The raphe valve has striae with uniseriate areolae continuing across a pronounced submarginal hyaline rim to the edge of the valve. The sternum valve has uniseriate dash-like areolae continuously from the valve face until the valve edge. Micromorphologically, these species possess two different open valvocopulae: only the raphe valvocopula has fimbriae; the sternum valvocopula has none. Based on p-distances of currently available DNA sequence data, i.e., rbcL and 18SV4, both species are pronouncedly different from the epitype strain of C. placentula, with C. coreana closest to the published molecular data of the strain UTEX FD23 named C. placentula from Iowa, USA, while C. sijunghoensis is closest but not the same as the publ is hed molecular data of strain D36_012, the epitype strain of C. placentula from Berlin, Germany. Based on scanning elect ron microscope observations, differentiating features are discussed concerning valvocopula fimbriae, central area, areolation of the sternum valve and on the raphe valve especially between the submarginal hyaline rim and edge

Exploring diatom diversity through cultures - a case study from the Bow River, Canada

Diatom cultures can help answer taxonomic, biogeographic and ecological questions on a local and global scale. Unialgal cultures are derived from a single cell and provide abundant material for morphological and molecular analyses. The link between the historic morphological species concept and the molecular data is becoming increasingly important with the use of eDNA metabarcoding. Additionally, cultures provide insights into the life cycle of diatoms and thereby complement taxonomy and species ecology. In this study, we present an approach to extract benthic diatoms from an environmental sample to generate unialgal cultures. We explored diatom diversity in preserved assemblages and by culturing as many different taxa as possible from benthic freshwater samples taken on the same day from the Bow River in Calgary, Canada. With both methods we found a total of 221 different benthic diatom taxa, of which 182 were identified in the preserved diatom assemblages. Interestingly, an additional 39 taxa only appeared in the cultures. In total 129 strains were cultivated representing 71 different taxa. This study includes pictures of living cells demonstrating the additional merits of unialgal cultures, as they provide information on plastid details, auxospores and endosymbionts. Both, the identification of the diatom assemblages and the generation and identification of strains provide the foundation for additional water quality assessment tools, taxonomic insights and molecular references libraries

Antarctic benthic diatoms after 10 months of dark exposure: consequences for photosynthesis and cellular integrity

Antarctic algae are exposed to prolonged periods of extreme darkness due to polar night, and coverage by ice and snow can extend such dark conditions to up to 10 months. A major group of microalgae in benthic habitats of Antarctica are diatoms, which are key primary producers in these regions. However, the effects of extremely prolonged dark exposure on their photosynthesis, cellular ultrastructure, and cell integrity remain unknown. Here we show that five strains of Antarctic benthic diatoms exhibit an active photosynthetic apparatus despite 10 months of dark-exposure. This was shown by a steady effective quantum yield of photosystem II (Y[II]) upon light exposure for up to 2.5 months, suggesting that Antarctic diatoms do not rely on metabolically inactive resting cells to survive prolonged darkness. While limnic strains performed better than their marine counterparts, Y(II) recovery to values commonly observed in diatoms occurred after 4-5 months of light exposure in all strains, suggesting long recovering times. Dark exposure for 10 months dramatically reduced the chloroplast ultrastructure, thylakoid stacking, and led to a higher proportion of cells with compromised membranes than in light-adapted cells. However, photosynthetic oxygen production was readily measurable after darkness and strong photoinhibition only occurred at high light levels (>800 µmol photons m-2 s-1). Our data suggest that Antarctic benthic diatoms are well adapted to long dark periods. However, prolonged darkness for several months followed by only few months of light and another dark period may prevent them to regain their full photosynthetic potential due to long recovery times, which might compromise long-term population survival

Lipid degradation and photosynthetic traits after prolonged darkness in four Antarctic benthic diatoms, including the newly described species Planothidium wetzelii sp. nov.

In polar regions, the microphytobenthos has important ecological functions in shallow-water habitats, such as on top of coastal sediments. This community is dominated by benthic diatoms, which contribute significantly to primary production and biogeochemical cycling while also being an important component of polar food webs. Polar diatoms are able to cope with markedly changing light conditions and prolonged periods of darkness during the polar night in Antarctica. However, the underlying mechanisms are poorly understood. In this study, five strains of Antarctic benthic diatoms were isolated in the field, and the resulting unialgal cultures were identified as four distinct species, of which one is described as a new species, Planothidium wetzelii sp. nov. All four species were thoroughly examined using physiological, cell biological, and biochemical methods over a fully controlled dark period of 3 months. The results showed that the utilization of storage lipids is one of the key mechanisms in Antarctic benthic diatoms to survive the polar night, although different fatty acids were involved in the investigated taxa. In all tested species, the storage lipid content declined significantly, along with an ultrastructurally observable degradation of the chloroplasts. Surprisingly, photosynthetic performance did not change significantly despite chloroplasts decreasing in thylakoid membranes and an increased number of plastoglobules. Thus, a combination of biochemical and cell biological mechanisms allows Antarctic benthic diatoms to survive the polar night

Exploring benthic diatom diversity in the West Antarctic Peninsula: insights from a morphological and molecular approach

Polar regions are among the most extreme habitats on Earth. However, diatom biodiversity in those regions is much more extensive and ecologically diverse than previously thought. The objective of this study was to add knowledge to benthic diatom biodiversity in Western Antarctic coastal zones via identification by means of morphology, DNA metabarcoding and cultured isolates. In addition, a taxonomically validated reference library for Antarctic benthic diatoms was established with comprehensive information on habitat, morphology and DNA barcodes (rbcL and 18SV4). Benthic samples from marine, brackish and freshwater habitats were taken at the Antarctic Peninsula. A total of 162 clonal cultures were established, resulting in the identification of 60 taxa. The combination of total morphological richness of 174 taxa, including the clones, with an additional 73 taxa just assigned by metabarcoding resulted in 247 infrageneric taxa. Of those taxa, 33 were retrieved by all three methods and 111 only by morphology. The barcode reference library of Antarctic species with the new references obtained through culturing allowed the assignment of 47 taxa in the metabarcoding analyses, which would have been left unassigned because no matching reference sequences were available before. Non–metric multidimensional scaling analyses of morphological as well as molecular data showed a clear separation of diatom communities according to water and substratum types. Many species, especially marine taxa, still have no record in reference databases. This highlights the need for a more comprehensive reference library to further improve routine diatom metabarcoding. Overall, a combination of morphological and molecular methods, along with culturing, provides complementary information on the biodiversity of benthic diatoms in the region

Antarctic benthic diatoms after 10 months of dark exposure: consequences for photosynthesis and cellular integrity

Antarctic algae are exposed to prolonged periods of extreme darkness due to polar night, and coverage by ice and snow can extend such dark conditions to up to 10 months. A major group of microalgae in benthic habitats of Antarctica are diatoms, which are key primary producers in these regions. However, the effects of extremely prolonged dark exposure on their photosynthesis, cellular ultrastructure, and cell integrity remain unknown. Here we show that five strains of Antarctic benthic diatoms exhibit an active photosynthetic apparatus despite 10 months of dark-exposure. This was shown by a steady effective quantum yield of photosystem II (Y[II]) upon light exposure for up to 2.5 months, suggesting that Antarctic diatoms do not rely on metabolically inactive resting cells to survive prolonged darkness. While limnic strains performed better than their marine counterparts, Y(II) recovery to values commonly observed in diatoms occurred after 4-5 months of light exposure in all strains, suggesting long recovering times. Dark exposure for 10 months dramatically reduced the chloroplast ultrastructure, thylakoid stacking, and led to a higher proportion of cells with compromised membranes than in light-adapted cells. However, photosynthetic oxygen production was readily measurable after darkness and strong photoinhibition only occurred at high light levels (>800 µmol photons m-2 s-1). Our data suggest that Antarctic benthic diatoms are well adapted to long dark periods. However, prolonged darkness for several months followed by only few months of light and another dark period may prevent them to regain their full photosynthetic potential due to long recovery times, which might compromise long-term population survival

Gomphonella olivacea (Bacillariophyceae) – a new phylogenetic position for a well-known taxon, its typification, new species and combinations

Background and aims – Within the project “German Barcode of Life – Diatoms” common diatoms of German waters were routinely isolated and cultivated. In order to understand the taxonomy and phylogeny of the genus Gomphonema, one of the most common taxa of Central Europe, known currently either under the name Gomphonema olivaceum in Europe or Gomphoneis olivacea in America, was studied. Methods – Twenty unialgal strains were established from five different water bodies in Germany and one from Lake Balaton, Hungary, which supplied molecular data (18S V4 and rbcL) besides morphometric and ultrastructural data. In addition, on eight populations from different water bodies including the type from Denmark, morphometric and micromorphological studies by light and scanning electron microscopy were performed. Key results – Molecular and micromorphological data show that the target taxon neither belongs to Gomphonema Ehrenb. nor to Gomphoneis Cleve. By reinstating the genus name Gomphonella Rabenh., the nomenclatural and taxonomic enigma of this taxon is solved, and with the presentation of the type by Hornemann the authorship of the epithet is clarified. Molecular data for the unialgal strains and several environmental clones show that there is more diversity in the Gomphonella olivacea clade than can be identified morphologically. In addition, the establishment of the new species Gomphonella coxiae and Gomphonella acsiae is supported. The molecular data classified Gomphonella species as belonging to the Cymbellales but not to the Gomphonemataceae. In addition, molecular data put Gomphoneis tegelensis R.Jahn & N.Abarca also into Gomphonella. In order to make the genera Gomphoneis and Gomphonema monophyletic, their astigmate members are transferred to Gomphonella. Conclusions – The results clarify that the gomphonemoid outline is not restricted to the family Gomphonemataceae but seem to be distributed across the entire order Cymbellales. This is shown in this paper for the revived genus Gomphonella, which contains the astigmate group of Gomphoneis and Gomphonema besides the longly disputed G. olivacea. Only a polyphasic approach, combining molecular and micromorphological data for taxonomy, nomenclatural evaluation, and observations from clonal cultures can reveal the full intricacies of evolutionary relations

Exploring benthic diatom diversity in the West Antarctic Peninsula: insights from a morphological and molecular approach

Polar regions are among the most extreme habitats on Earth. However, diatom biodiversity in those regions is much more extensive and ecologically diverse than previously thought. The objective of this study was to add knowledge to benthic diatom biodiversity in Western Antarctic coastal zones via identification by means of morphology, DNA metabarcoding and cultured isolates. In addition, a taxonomically validated reference library for Antarctic benthic diatoms was established with comprehensive information on habitat, morphology and DNA barcodes (rbcL and 18SV4). Benthic samples from marine, brackish and freshwater habitats were taken at the Antarctic Peninsula. A total of 162 clonal cultures were established, resulting in the identification of 60 taxa. The combination of total morphological richness of 174 taxa, including the clones, with an additional 73 taxa just assigned by metabarcoding resulted in 247 infrageneric taxa. Of those taxa, 33 were retrieved by all three methods and 111 only by morphology. The barcode reference library of Antarctic species with the new references obtained through culturing allowed the assignment of 47 taxa in the metabarcoding analyses, which would have been left unassigned because no matching reference sequences were available before. Non–metric multidimensional scaling analyses of morphological as well as molecular data showed a clear separation of diatom communities according to water and substratum types. Many species, especially marine taxa, still have no record in reference databases. This highlights the need for a more comprehensive reference library to further improve routine diatom metabarcoding. Overall, a combination of morphological and molecular methods, along with culturing, provides complementary information on the biodiversity of benthic diatoms in the region

MMX Rover Locomotion Subsystem - Development and Testing towards the Flight Model

Wheeled rovers have been successfully used as mobile landers on Mars and Moon and more such missions are in the planning. For the Martian Moon eXploration (MMX) mission of the Japan Aerospace Exploration Agency (JAXA), such a wheeled rover will be used on the Marsian Moon Phobos. This is the first rover that will be used under such low gravity, called milli-g, which imposes many challenges to the design of the locomotion subsystem (LSS). The LSS is used for unfolding, standing up, driving, aligning and lowering the rover on Phobos. It is a entirely new developed highly-integrated mechatronic system that is specifically designed for Phobos. Since the Phase A concept of the LSS, which was presented two years ago [1], a lot of testing, optimization and design improvements have been done. Following the tight mission schedule, the LSS qualification and flight models (QM and FM) assembly has started in Summer 2021. In this work, the final FM design is presented together with selected test and optimization results that led to the final state. More specifically, advances in the mechanics, electronics, thermal, sensor, firmware and software design are presented. The LSS QM and FM will undergo a comprehensive qualification and acceptance testing campaign, respectively, in the first half of 2022 before the FM will be integrated into the rove