Internal Transcribed Spacer and 16S Amplicon Sequencing Identifies Microbial Species Associated with Asbestos in New Zealand.

Inhalation of asbestos fibres can cause lung inflammation and the later development of asbestosis, lung cancer, and mesothelioma, and the use of asbestos is banned in many countries. In most countries, large amounts of asbestos exists within building stock, buried in landfills, and in contaminated soil. Mechanical, thermal, and chemical treatment options do exist, but these are expensive, and they are not effective for contaminated soil, where only small numbers of asbestos fibres may be present in a large volume of soil. Research has been underway for the last 20 years into the potential use of microbial action to remove iron and other metal cations from the surface of asbestos fibres to reduce their toxicity. To access sufficient iron for metabolism, many bacteria and fungi produce organic acids, or iron-chelating siderophores, and in a growing number of experiments these have been found to degrade asbestos fibres in vitro. This paper uses the internal transcribed spacer (ITS) and 16S amplicon sequencing to investigate the fungal and bacterial diversity found on naturally-occurring asbestos minerals, asbestos-containing building materials, and asbestos-contaminated soils with a view to later selectively culturing promising species, screening them for siderophore production, and testing them with asbestos fibres in vitro. After filtering, 895 ITS and 1265 16S amplicon sequencing variants (ASVs) were detected across the 38 samples, corresponding to a range of fungal, bacteria, cyanobacterial, and lichenized fungal species. Samples from Auckland (North Island, New Zealand) asbestos cement, Auckland asbestos-contaminated soils, and raw asbestos rocks from Kahurangi National Park (South Island, New Zealand) were comprised of very different microbial communities. Five of the fungal species detected in this study are known to produce siderophores.

Chemical diversity of kanuka: Inter- and intraspecific variation of foliage terpenes and flavanones of Kunzea (Myrtaceae) in Aotearoa/New Zealand.

Kunzea (Myrtaceae) trees and shrubs, generally called kanuka, grow across most of Aotearoa/New Zealand (NZ). With the exception of K. sinclairii, an offshore island endemic, kanuka had been treated as an Australasian species K. ericoides. However, a 2014 taxonomic revision recognized ten species, all endemic to NZ. Kanuka chemistry is less studied than that of its closest relative in NZ, manuka (Leptospermum scoparium), which shows very distinct regional foliage chemotypes. We have used a miniaturized method with GC and 1H NMR to analyze foliage chemistry of voucher specimens from across the geographic ranges of the ten NZ Kunzea species. We found common mono- and sesquiterpenes, with α-pinene dominant in all samples, but only traces of antimicrobial triketones. Two unusual flavanones, with unsubstituted B-rings and known bioactivity against Phytophthora, did distinguish some of the samples. 5,7-Dihydroxy-6,8-dimethyl flavanone was only found at high concentrations in the three K. sinclairii samples in this study's sample set, but this compound has separately been reported in K. robusta samples from a nearby region. Therefore none of the NZ Kunzea species was distinguished by the chemistry analyzed in this study, but there is a possibility of regional flavonoid chemotypes cutting across the species boundaries.

Chatbots as a Tool to Scale Mentoring Processes: Individually Supporting Self-Study in Higher Education.

Like most curricula in the humanities and social sciences, the curriculum of pre-service teacher training in educational sciences often includes time-consuming reading and writing tasks, which require high quality support and feedback in a timely manner. A well-known way to provide this support to students is one-to-one mentoring. However, limited time and resources in the German university context require to effectively scale the benefits of individual feedback. The use of scalable technologies to support learning processes seems to be promising, but its development usually requires a deep technical understanding. With an interdisciplinary approach, this contribution investigates how personal mentoring can be made available to as many students as possible, taking into account the didactic, organizational and technical frameworks at universities. We describe the development and implementation process of two chatbots that both aim to support students of educational sciences in their self-study of the seminar topics and literature. The chatbots were used by over 700 students during the course of 1 year and our evaluations show promising results that bear the potential to improve the availability of digital mentoring support for all students.

The indigenous vascular flora of the forest domain of Anela (Sardinia, Italy).

The importance of mountains for plant diversity and richness is underestimated, particularly when transition zones between different bioclimates are present along altitudinal gradients. Here we present the first floristic data for a mountain area in the island of Sardinia (Italy), which exhibits Mediterranean bioclimates at the bottom and temperate bioclimate at the top. We discovered a very high floristic richness, despite the fact that the number of endemic taxa is not high and the number of exclusive taxa is very low. Many of the detected taxa are at their range periphery and/or ecological margin. We conclude that climate transition zones in Mediterranean mountains and especially on islands are key areas regarding plant biodiversity and should be better investigated and protected.

Genetic structuring of the coastal herb Arthropodium cirratum (Asparagaceae) is shaped by low gene flow, hybridization and prehistoric translocation.

We examined the genetic structuring of rengarenga (Arthropodium cirratum; Asparagaceae), an endemic New Zealand coastal herb, using nuclear microsatellite markers. This species was brought into cultivation by Maori within the last 700-800 years for its edible roots and was transplanted beyond its natural distribution as part of its cultivation. We found very high levels of genetic structuring in the natural populations (FST = 0.84), indicating low levels of gene flow. Reduced genetic diversity was found in the translocated populations, suggesting a large loss of genetic diversity early in the domestication process. The data indicates that rengarenga was brought into cultivation independently at least three times, with the sources of these introductions located within a narrow area encompassing about 250km of coastline. Hybridization was inferred between A. cirratum and the closely related A. bifurcatum, despite A. birfucatum not occurring in the vicinity.

Anemonastrum tenuicaule and A. antucense (Ranunculaceae), new combinations for a New Zealand endemic species and its South American relative.

A rational taxonomic circumscription of genera in tribe Anemoneae (Ranunculaceae) is briefly discussed. It is concluded that, in view of the morphological diversity of the group and recent molecular phylogenetic findings, a moderately narrow approach to the re-circumscription of genera earlier included in Anemone sensu lato is preferable, in particular, with the recognition of the lineage with the base chromosome number x = 7 (Anemone subgen. Anemonidium) as two genera, Hepatica sensu stricto and Anemonastrum in an expanded circumscription (including Anemonidium, Arsenjevia, Jurtsevia, and Tamuria). Following these conclusions, new nomenclatural combinations are proposed for two related species endemic to New Zealand and South America, respectively: Anemonastrum tenuicaule (= Anemone tenuicaulis, Ranunculus tenuicaulis) and Anemonastrum antucense (= Anemone antucensis). Information on typification is updated: the lectotype of Anemone antucensis is the specimen from P and not a specimen from G, and the lectotype of Ranunculus tenuicaulis is a specimen from AK. Biogeographic scenarios already proposed to explain the relationship of these two species and some other South America - New Zealand distribution patterns are discussed. It is concluded that the long-distance dispersal scenario fits best the available data for Anemonastrum. Two host-specific and geographically restricted species of Urosystis parasitizing A. tenuicaule and A. antucense are briefly discussed.

Using citizen science to bridge taxonomic discovery with education and outreach.

PREMISE OF THE STUDY: Biological collections are uniquely poised to inform the stewardship of life on Earth in a time of cataclysmic biodiversity loss. Efforts to fully leverage collections are impeded by a lack of trained taxonomists and a lack of interest and engagement by the public. We provide a model of a crowd-sourced data collection project that produces quality taxonomic data sets and empowers citizen scientists through real contributions to science. Entitled MicroPlants, the project is a collaboration between taxonomists, citizen science experts, and teachers and students from universities and K-12. METHODS: We developed an online tool that allows citizen scientists to measure photographs of specimens of a hyper-diverse group of liverworts from a biodiversity hotspot. RESULTS: Using the MicroPlants online tool, citizen scientists are generating high-quality data, with preliminary analysis indicating non-expert data can be comparable to expert data. DISCUSSION: More than 11,000 users from both the website and kiosk versions have contributed to the data set, which is demonstrably aiding taxonomists working toward establishing conservation priorities within this group. MicroPlants provides opportunities for public participation in authentic science research. The project's educational component helps move youth toward engaging in scientific thinking and has been adopted by several universities into curriculum for both biology and non-biology majors.

Isolation and characterization of microsatellite loci from Arthropodium cirratum (Asparagaceae).

PREMISE OF THE STUDY: Microsatellite markers were developed for Arthropodium cirratum (Asparagaceae) to study population genetic structure and translocation of this species. These markers were tested for cross-amplification in two other Arthropodium species. METHODS AND RESULTS: Sixteen microsatellite markers were developed from a genomic library and tested in three populations of A. cirratum. The loci exhibited one to five alleles per locus, with private alleles present in each of the populations. Cross-amplification tests in the two other New Zealand Arthropodium species revealed that many of the loci amplify and demonstrate polymorphism in A. bifurcatum. CONCLUSIONS: These markers will be useful for determining genetic structure in A. cirratum and for determining the origins of translocated populations of this species.

Species delimitation and biogeography of a southern hemisphere liverwort clade, Frullania subgenus Microfrullania (Frullaniaceae, Marchantiophyta).

Frullania subgenus Microfrullania is a clade of ca. 15 liverwort species occurring in Australasia, Malesia, and southern South America. We used combined nuclear and chloroplast sequence data from 265 ingroup accessions to test species circumscriptions and estimate the biogeographic history of the subgenus. With dense infra-specific sampling, we document an important role of long-distance dispersal in establishing phylogeographic patterns of extant species. At deeper time scales, a combination of phylogenetic analyses, divergence time estimation and ancestral range estimation were used to reject vicariance and to document the role of long-distance dispersal in explaining the evolution and biogeography of the clade across the southern Hemisphere. A backbone phylogeny for the subgenus is proposed, providing insight into evolution of morphological patterns and establishing the basis for an improved sectional classification of species within Microfrullania. Several species complexes are identified, the presence of two undescribed but genetically and morphologically distinct species is noted, and previously neglected names are discussed.

Evidence of a Strong Domestication Bottleneck in the Recently Cultivated New Zealand Endemic Root Crop, Arthropodium cirratum (Asparagaceae).

We use chloroplast DNA sequencing to examine aspects of the pre-European Maori cultivation of an endemic New Zealand root crop, Arthropodium cirratum (rengarenga). Researching the early stages of domestication is not possible for the majority of crops, because their cultivation began many thousands of years ago and/or they have been substantially altered by modern breeding methods. We found high levels of genetic variation and structuring characterised the natural distribution of A. cirratum, while the translocated populations only retained low levels of this diversity, indicating a strong bottleneck even at the early stages of this species' cultivation. The high structuring detected at four chloroplast loci within the natural A. cirratum range enabled the putative source(s) of the translocated populations to be identified as most likely located in the eastern Bay of Plenty/East Cape region. The high structuring within A. cirratum also has implications for the conservation of genetic diversity within this species, which has undergone recent declines in both its natural and translocated ranges.

Systematics of Simplicia Kirk (Poaceae, Agrostidinae) - an endemic, threatened New Zealand grass genus.

A new species of the New Zealand endemic grass Simplicia, Simplicia felix is described. The new species is segregated from and compared with Simplicia buchananii and Simplicia laxa. Simplicia felix occurs mostly in lightly shaded areas of seasonally dry alluvial forest. A distribution map and an assessment of the conservation status of the new species are presented. Genetic variation in the genus was examined, building on previously published work but including additional sampling. Analysis of nrDNA ITS and ETS and plastid trnL intron and trnL-F intergenic spacer sequences show Simplicia felix to be more closely related to Simplicia laxa than to Simplicia buchananii. NeighborNet analyses of AFLP profiles for the three species of Simplicia show each to consist of distinct clusters of genotypes well separated from each other.

A revision of the New Zealand Kunzeaericoides (Myrtaceae) complex.

A revision of the New Zealand Kunzeaericoides complex is presented. This paper is the final of a series that has explored the systematics of the New Zealand Kunzea complex using cytological and molecular variation, as well as experimental hybridisations between postulated segregates. As a result of those studies ten species, all endemic to New Zealand, are recognised; seven of these are new. One species, Kunzeatriregensis sp. nov., is endemic to the Three Kings Islands and another species Kunzeasinclairii, endemic to Aotea (Great Barrier Island). The North Island of New Zealand has seven species, Kunzeaamathicola sp. nov., Kunzeasalterae sp. nov., Kunzeaserotina sp. nov., Kunzearobusta sp. nov., Kunzeatenuicaulis sp. nov., Kunzeatoelkenii sp. nov., and Kunzealinearis comb. nov. Of these, Kunzealinearis, Kunzeasalterae, Kunzeatenuicaulis and Kunzeatoelkenii are endemic to the North Island, and Kunzeaamathicola, Kunzearobusta and Kunzeaserotina extend to the South Island which also supports one endemic, Kunzeaericoides. Typifications are published for Leptospermumericoides A.Rich., Leptospermumericoidesvar.linearis Kirk, Leptospermumericoidesvar.microflorum G.Simps., Leptospermumericoidesvar.pubescens Kirk, and Leptospermumsinclairii Kirk, names here all referred to Kunzea. The ecology, conservation, extent of natural hybridisation and some aspects of the ethnobotany (vernacular names) of these Kunzea are also discussed.

Species tree phylogeny and character evolution in the genus Centipeda (Asteraceae): evidence from DNA sequences from coding and non-coding loci from the plastid and nuclear genomes.

A species tree phylogeny of the Australian/New Zealand genus Centipeda (Asteraceae) is estimated based on nucleotide sequence data. We analysed sequences of nuclear ribosomal DNA (ETS, ITS) and three plasmid loci (ndhF, psbA-trnH, and trnL-F) using the multi-species coalescent module in BEAST. A total of 129 individuals from all 10 recognised species of Centipeda were sampled throughout the species distribution ranges, including two subspecies. We conclude that the inferred species tree topology largely conform previous assumptions on species relationships. Centipeda racemosa (Snuffweed) is the sister to remaining species, which is also the only consistently perennial representative in the genus. Centipeda pleiocephala (Tall Sneezeweed) and C. nidiformis (Cotton Sneezeweed) constitute a species pair, as does C. borealis and C. minima (Spreading Sneezeweed), all sharing the symplesiomorphic characters of spherical capitulum and convex receptacle with C. racemosa. Another species group comprising C. thespidioides (Desert Sneezeweed), C. cunninghamii (Old man weed, or Common sneeze-weed), C. crateriformis is well-supported but then include the morphologically aberrant C. aotearoana, all sharing the character of having capitula that mature more slowly relative the subtending shoot. Centipeda elatinoides takes on a weakly supported intermediate position between the two mentioned groups, and is difficult to relate to any of the former groups based on morphological characters.

Frullania knightbridgei, a new liverwort (Frullaniaceae, Marchantiophyta) species from the deep south of Aotearoa-New Zealand based on an integrated evidence-based approach.

Frullania is a large and taxonomically complex genus. A new liverwort species, Frullania knightbridgeisp. nov. from southern New Zealand, is described and illustrated. The new species, and its placement in Frullania subg. Microfrullania, is based on an integrated evidence-based approach derived from morphology, ecology, experimental growth studies of plasticity, as well as sequence data. Diagnostic characters associated with the leaf and lobule cell-wall anatomy, oil bodies, and spore ultra-structure distinguish it from all other New Zealand species of Frullania. A critical comparison is also made between Frullania knightbridgei and morphologically allied species of botanical regions outside the New Zealand region and an artificial key is provided. The new species is similar to some forms of the widespread Australasian species, Frullania rostrata, but has unique characters associated with the lobule and oil bodies. Frullania knightbridgei is remarkably interesting in comparison with the majority of Frullania species, and indeed liverworts in general, in that it is at least partially halotolerant. Maximum parsimony and maximum likelihood analyses of nuclear ribosomal ITS2 and plastidic trnL-trnF sequences from purported related speciesconfirms its independent taxonomic status and corroborates its placement within Frullania subg. Microfrullania.

Parallel polyploid speciation: distinct sympatric gene-pools of recurrently derived allo-octoploid Asplenium ferns.

Although polyploidy is widespread, its significance to the generation of biodiversity remains unclear. Many polyploids have been derived recurrently. For a particular polyploid, gene-flow between the products of independent origin is typical where they come into contact. Here, we use AFLP DNA-fingerprinting and chloroplast DNA sequences to demonstrate parallel polyploid speciation within both of the ferns Asplenium cimmeriorum and A. gracillimum. Both of these taxa comprise at least two allopolyploids, recurrently derived from the same progenitor pair. Each of these allopolyploids remain genetically distinguishable even with extensive sympatry, and could therefore be considered distinct species. To our knowledge, parallel speciation on this scale amongst recurrent polyploids has not been previously reported. With their parallel origins, these 'evolutionary replicates' provide an unrivalled opportunity to investigate how the reproductive barriers and ecological differentiation necessary for speciation arise following polyploidy.

Chemotaxonomy of Pseudowintera: sesquiterpene dialdehyde variants are species markers.

Two sesquiterpene dialdehydes, the 1beta-E-coumaroyl-5alpha-hydroxypolygodial plus the known 1beta-E-cinnamoyl-5alpha-hydroxypolygodial, were isolated from the recently described species Pseudowintera insperata. This discovery is a further example of the rare sesquiterpene dialdehyde coumarate/cinnamate combination being found exclusively in the family Winteraceae. Another sesquiterpene dialdehyde, isopaxidal, with the rare rearranged drimane skeleton, was isolated from Pseudowintera axillaris. The sesquiterpene dialdehyde contents of leaves of 25 individual plants of the four Pseudowintera species, all endemic to New Zealand, were measured by HPLC. P. insperata individuals all had high levels (3.0-6.9% of leaf dry wt.) of the coumarate, P. axillaris had high levels (2.2-6.9%) of paxidal, and Pseudowintera colorata from different areas of New Zealand contained varying levels of polygodial (1.4-2.9%) and 9-deoxymuzigadial (0-2.9%). Therefore the sesquiterpene dialdehydes are good species markers.

Difference in pathogenesis between vitiligo vulgaris and halo nevi associated with vitiligo is supported by an HLA association study.

Human leukocyte antigen (HLA) class II associations with two subtypes of vitiligo: vitiligo vulgaris and halo nevi associated with vitiligo were investigated. In previous studies associations between vitiligo and HLA antigens have been reported but these two subtypes have never been taken into account. However from a clinical and histological point of view, a difference in (auto)-immune pathogenesis can be expected. This difference might be reflected in an association with different HLA alleles. Seventy-six unrelated Dutch Caucasians, 40 with vitiligo vulgaris and 36 with halo nevi associated with vitiligo were included. A panel of randomly chosen HLA typed healthy Dutch blood donors (n = 2400) served as control population. HLA-DR and -DQ typing was carried out on blood samples by amplifying genomic DNA using polymerase chain reaction followed by dot blot hybridization with sequence specific oligonucleotides. The main outcome measures were odds ratio (OR), uncorrected P-value (P(u)) and corrected P-value. There were distinct differences in the clinical manifestations between vitiligo vulgaris and halo nevi associated with vitiligo with respect to precipitating factors, extent and progress of the disease and the association with other auto-immune diseases in the two subtypes and their respective first degree family members. Our stratification reveals differences in HLA class II between both subtypes and between subtypes and controls. A case-control association study showed a significant positive association of HLA-DR4 (OR = 2.787, P(u) = 0.0022) and DR53 (OR = 2.249, P(u) = 0.0153) and a negative association of HLA-DR3 (OR = 0.195, P(u) = 0.0024) with vitiligo vulgaris. The group with halo nevi associated with vitiligo did not show these associations, but had a significant negative association with HLA-DR11 (OR = 0.083, P(u) = 0.0067). In conclusion, the differences in HLA association within clinical subtypes of vitiligo support our suggestion that vitiligo vulgaris and halo nevi associated with vitiligo have distinct pathogenic mechanisms.

The number of amino acid triplet differences between patient and donor is predictive for the antibody reactivity against mismatched human leukocyte antigens.

BACKGROUND: The correlation between antibody production against mismatched donor human leukocyte antigens (HLA) and the number of amino acid sequence mismatches was analyzed in patients who rejected a kidney transplant (n=146). METHODS: A similar analysis was performed for the antibody production of women against the paternal HLA antigens of their child (n=1,397). The amino acid sequence (triplet) differences were analyzed using the HLAMatchmaker algorithm. RESULTS: In both groups, a positive correlation was found between the number of triplet mismatches and the percentage of individuals producing antibodies (P <0.0001). If zero triplet mismatches were present, no antibodies were formed in all cases. When 11 or 12 triplet mismatches were present, 94% of the transplant patients produced antibodies against the donor. In pregnancy, 11 or 12 triplet mismatches led to 27% of the women producing specific antibodies. CONCLUSIONS: These results indicate that the immunogenicity of the fetus is lower than that of a rejected kidney and that analysis of the number of triplet mismatches can predict the antibody reactivity against the mismatched HLA antigens.

The HLA-DR phenotype of the responder is predictive of humoral response against HLA class I antigens.

Recent studies suggest that the immunogenicity of an human leukocyte antigen (HLA) incompatibility should be considered in the context of the HLA phenotype of the recipient. The HLA-DR phenotype of the responder is thought to be predictive for the strength of the alloimmune response. In order to analyze the humoral response against HLA class I antigens in the context of the HLA-DR phenotype of the responder, we selected all HLA-DR homozygous Dutch patients that were present on the Eurotransplant waiting list between 1967 and 2000 (n=1,317 patients). By logistic regression it was determined whether antibody production against a specific HLA class I antigen is associated with a particular HLA-DR antigen in the patient. Furthermore, it was analyzed whether a patient, expressing a particular HLA-DR antigen, preferentially produces antibodies against particular HLA class I antigens. The results demonstrate that patients, homozygous for a certain HLA-DR antigen, cannot be considered high or low responders when analyzing the antibody response in terms of panel reactive antibody (PRA) value. However, a correlation can be found between the HLA-DR phenotype of the patient and the specific antibody response against HLA class I antigens. For example, antibodies against HLA-A10, -A11, -A19, and -B35 are produced more frequently by HLA-DR6 positive individuals, whereas antibodies against HLA-A3, -B5, -B7, -B8, and -B12 are produced more frequently by HLA-DR4 positive individuals. These data confirm that the HLA-DR phenotype of the responder plays a determinative role in the immunogenicity of mismatched HLA antigens. The results indicate that selection of HLA class I mismatches of the donor in the context of the HLA-DR phenotype of the responder might reduce the incidence of humoral graft rejection and minimize the sensitization grade of retransplant candidates.

Differential immunogenicity of paternal HLA Class I antigens in pregnant women.

More insight into the differential immunogenicity of human leukocyte antigen (HLA) mismatches will be beneficial for donor selection in clinical transplantation. In this study the immunogenicity of HLA antigens was analyzed by examining the antibody profiles in women who have been pregnant. In total 888 women, who had pregnancy induced HLA alloantibodies, were included in this study, while 413 women who had not been immunized by their pregnancy, served as controls. First it was analyzed whether women expressing particular HLA antigens are more likely to produce HLA alloantibodies. Next we determined whether certain HLA mismatches of their children are more immunogenic than other ones. Finally we studied whether the immunogenicity of specific HLA mismatches is dependent on the HLA phenotype of the women. Women expressing HLA-A3, HLA-A32, and HLA-B21 are more likely to produce alloantibodies whereas women expressing HLA-B13 and HLA-B17 have a significantly lower incidence of alloantibodies compared with women expressing other HLA antigens. Children with HLA-A2 or HLA-B5 mismatches induced alloantibodies significantly more often whereas children with HLA-A30, -A31 or -A33 and HLA-A28 induced alloantibodies significantly less often than children with other HLA class I mismatches. Finally we could demonstrate that the immunogenicity of a particular HLA mismatch is dependent on the HLA phenotype of the women. Information on the differential immunogenicity of HLA mismatches may be of benefit for the determination of acceptable and taboo mismatches in the case of donor selection for (highly sensitized) patients.