Research@Lincoln
Research@Lincoln is an open access institutional repository collecting the research produced by Lincoln University staff and students. You may also be interested in Data@Lincoln or Lincoln University Living Heritage.
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Publication Open Access The abundance, activity, and community composition of comammox Nitrospira and canonical ammonia oxidisers in New Zealand soils : A thesis submitted in partial fulfilment of the requirements for the Degree of Doctor of Philosophy at Lincoln University(Lincoln University, 2024)Nitrification, the microbial oxidation of ammonia (NH3) via nitrite (NO2-) to nitrate (NO3-) is an important process in terrestrial ecosystems, as it contributes to the production of two environmentally significant products, nitrous oxide (N2O), and nitrate. Traditionally, nitrification was thought to be a two-step process, where ammonia is first converted to nitrite by ammonia-oxidising bacteria (AOB) before nitrite oxidising bacteria (NOB) complete the oxidation to nitrate. Later, it was discovered that a group of archaea (ammonia oxidising archaea, AOA) could also undertake ammonia oxidation, typically in oligotrophic/extreme conditions such as low ammonia availability and pH. The separation of nitrification into two steps involving different microorganisms has puzzled scientists as a single organism completing both steps of nitrification was theoretically assumed to be more efficient. The presence of complete ammonia oxidisers (comammox) was later confirmed by cultivating them from an aquaculture system and a core from a deep-sea oil well. It was found that comammox belongs to lineage II of the genus Nitrospira, a group of bacteria traditionally thought to be responsible for nitrite oxidation. Comammox Nitrospira and canonical Nitrospira can be distinguished by the presence of the ammonia monooxygenase gene (AMO). Furthermore, comammox Nitrospira can be separated into clade A and clade B based on the phylogeny of this gene. Clade A can also be further divided into sub-clades A.1, A.2.1, A2.2, and A.3. Since its initial discovery, comammox Nitrospira has been found in a variety of terrestrial ecosystems. Typically, studies have shown that clade B are more abundant in forest and paddy soils, whilst clade A.2 may prefer agricultural soils. Clade A.1 is seen to be the dominant cluster in natural and artificial aquatic ecosystems such as freshwater wells and wastewater treatment plants. However, the presence and distribution of comammox Nitrospira, relative to canonical ammonia oxidisers, in different soils and relationships with soil and environmental conditions are not fully understood. The research described in this Thesis was designed to fill this knowledge gap and improve our understanding of comammox Nitrospira and canonical ammonia oxidisers. Experiment 1 determined the abundance and community composition of comammox Nitrospira throughout New Zealand Dairy farms and quantified the abundance and community composition of comammox Nitrospira in New Zealand under various land uses. It was concluded that comammox Nitrospira are ubiquitous throughout New Zealand soils. Comammox Nitrospira amoA abundance shared a strong positive and strong negative correlation with soil moisture and pH, respectively. Interestingly, the sequencing analysis determined that the comammox Nitrospira community solely consisted of clade B. Two experiments were devised to investigate these correlations, with respect to canonical ammonia oxidisers. Experiment 2 explored the effect of soil pH, with N inputs, on comammox Nitrospira abundance and community composition in New Zealand dairy pasture soils. Comammox Nitrospira preferred the natural (6.1-6.2) soil pH with no nitrogen amendment. Comparatively, the AOB community (dominated by Nitrosospira) responded positively to soil pH and nitrogen input. This may be due to the difference in ammonia availability. Estimated ammonia availability in the synthetic urine treatments (equivalent to 700 kg N ha-1, N700) accurately predicted the AOB amoA gene abundance. Interestingly, the AOA communities (which were predominantly made up of Thaumarchaeota group I.1b clade E) seemed to prefer the natural and high pH soils over the low pH. This may be due to the lineage of AOA present. AOA did not respond to the application of nitrogen. Experiment 3 investigated the effect of soil moisture and temperature on comammox Nitrospira abundance, transcriptional activity, and community composition, relative to canonical ammonia oxidisers. AOB was the dominant nitrifier in the synthetic urine treated soil regardless of temperature or moisture. Peak AOB amoA transcript abundance was positively correlated with estimated soil ammonia availability. While the nitrification rate and changes in AOB amoA gene abundance followed a similar relationship. Ammonia oxidising archaea were strongly influenced by soil temperature. At 20 °C, AOA amoA peak transcript abundance averaged over 1 order of magnitude higher than at 8 °C. A member of the AOA community associated with the Nitrosocosmicus subclade was positively correlated with ammonium and estimated soil ammonia concentrations. The presence and relative increase of Nitrosocosmicus AOA in a high nitrogen environment poses an interesting contrast to the current scientific opinion. Contrary to Experiment 1, the abundance of comammox Nitrospira amoA was not positively correlated with soil moisture. This suggests that the association is more complex than previously thought. Further research is required to determine the drivers of comammox Nitrospira abundance in a high moisture environment. Overall, the results of this thesis indicate that in New Zealand, AOB are the dominant ammonia oxidiser in a nitrogen-rich environment, such as a dairy farm soil. While the majority of the AOA community prefer a high temperature, low nitrogen environment. However, Nitrosocosmicus-like AOA may respond positively to nitrogen amendment, which challenges our current understanding of terrestrial AOA. Comammox Nitrospira may prefer a slightly acidic, oligotrophic soil environment and do not respond to temperature change. However, they may be the main ammonia oxidiser in some high moisture environments, potentially due to biotic interactions with plants or microbes.Item Restricted Trends in DNA barcoding and metabarcoding(NRC Research Press, 2019-03)This open-access special issue features 12 full articles representing emerging trends from the international DNA barcoding community. Several articles highlight how DNA-based techniques are elucidating the species diversity, biogeography, and conservation status of Africa’s biodiversity. Another prominent theme is the movement towards big biodiversity data using high-throughput, individual-based DNA barcoding methods, which preserve voucher specimens and abundance data, as well as bulk sample-based metabarcoding. Methodological developments are enhancing the detection of specific species and whole communities using environmental DNA (eDNA) barcoding and metabarcoding. Data are also expanding in terms of genetic coverage; in this issue, a new database is established for a secondary fungal DNA barcode marker, and multi-kingdom, multi-marker biodiversity surveys are gaining traction. DNA barcode sequence data, often combined with complementary markers or taxonomic information, are increasingly contributing to large-scale phylogenetic projects, with implications for understanding evolutionary history, community structure, and conservation priorities.Item Restricted Metagenomic insights to the functional potential of sediment microbial communities in freshwater lakes(Pensoft Publishers, 2022-03-25)Molecular-based techniques offer considerable potential to provide new insights into the impact of anthropogenic stressors on lake ecosystems. Microbial communities are involved in many geochemical cycling processes in lakes and a greater understanding of their functions could assist in guiding more targeted remedial actions. Recent advances in metagenomics now make it possible to determine the functional potential of entire microbial communities. The present study investigated microbial communities and their functional potential in surface sediments collected from three lakes with differing trophic states and characteristics. Surface sediments were analysed for their nutrient and elemental contents and metagenomics and metabarcoding analysis undertaken. The nutrients content of the surface sediments did not show as distinct a gradient as water chemistry monitoring data, likely reflecting effects of other lake characteristics, in particular, depth. Metabarcoding and metagenomics revealed differing bacterial community composition and functional potential amongst lakes. Amongst the differentially abundant metabolic pathways, the most prominent were clusters in the energy and xenobiotics pathways. Differences in the energy metabolism paths of photosynthesis and oxidative phosphorylation were observed. These were most likely related to changes in the community composition and especially the presence of cyanobacteria in two of the three lakes. Xenobiotic pathways, such as those involving polycyclic aromatic hydrocarbons, were highest in the lakes with the greatest agricultural land-use in their catchment. These results highlight how microbial metagenomics can be used to gain insights into the causes of differences in trophic status amongst lakes.Item Restricted Tetrodotoxin in marine bivalves and edible gastropods: A mini-review(Elsevier, 2019-12)Tetrodotoxin (TTX) is a potent neurotoxin responsible for countless human intoxications and deaths around the world. The distribution of TTX and its analogues is diverse and the toxin has been detected in organisms from both marine and terrestrial environments. Increasing detections seafood species, such as bivalves and gastropods, has drawn attention to the toxin, reinvigorating scientific interest and regulatory concerns. There have been reports of TTX in 21 species of bivalves and edible gastropods from ten countries since the 1980's. While TTX is structurally dissimilar to saxitoxin (STX), another neurotoxin detected in seafood, it has similar sodium channel blocking action and potency and both neurotoxins have been shown to have additive toxicities. The global regulatory level for the STX group toxins applied to shellfish is 800 μg/kg. The presence of TTX in shellfish is only regulated in one country; The Netherlands, with a regulatory level of 44 μg/kg. Due to the recent interest surrounding TTX in bivalves, the European Food Safety Authority established a panel to assess the risk and regulation of TTX in bivalves, and their final opinion was that a concentration below 44 μg of TTX per kg of shellfish would not result in adverse human effects. In this article, we review current knowledge on worldwide TTX levels in edible gastropods and bivalves over the last four decades, the different methods of detection used, and the current regulatory status. We suggest research needs that will assist with knowledge gaps and ultimately allow development of robust monitoring and management protocols.Item Restricted Seasonal and spatial variations in bacterial communities from tetrodotoxin-bearing and non-tetrodotoxin-bearing clams(Frontiers Media S.A., 2020-08-05)Tetrodotoxin (TTX) is one of the most potent naturally occurring compounds and is responsible for many human intoxications worldwide. Paphies australis are endemic clams to New Zealand which contain varying concentrations of TTX. Research suggests that P. australis accumulate the toxin exogenously, The aim of this study was to identify potential bacterial TTX-producers by exploring differences in bacterial communities in two organs of P. australis: the siphon and digestive gland. Samples from the digestive glands of a non-toxic bivalve Austrovenus stutchburyi that lives amongst toxic P. australis populations were also analyzed. Bacterial communities were characterized using 16S ribosomal RNA gene metabarcoding in P. australis sourced monthly from the Hokianga Harbor, a site known to have TTXbearing clams, for 1 year, from ten sites with varying TTX concentrations around New Zealand, and in A. stutchburyi from the Hokianga Harbor. Tetrodotoxin was detected in P. australis from sites all around New Zealand and in all P. australis collected monthly from the Hokianga Harbor. The toxin averaged 150 µg kg−1 over the year of sampling in the Hokianga Harbor but no TTX was detected in the A. stutchburyi samples from the same site. Bacterial species diversity differed amongst sites (p < 0.001, F = 5.9) and the diversity in siphon samples was significantly higher than in digestive glands (p < 0.001, F = 65.8). Spirochaetaceae (4–60%) and Mycoplasmataceae (16–78%) were the most abundant families in the siphons and the digestive glands, respectively. The bacterial communities were compared between sites with the lowest TTX concentrations and the Hokianga Harbor (site with the highest TTX concentrations), and the core bacterial communities from TTX-bearing individuals were analyzed. The results from both spatial and temporal studies corroborate with previous hypotheses that Vibrio and Bacillus could be responsible for the source of TTX in bivalves. The results from this study also indicate that marine cyanobacteria, in particular picocyanobacteria (e.g., Cyanobium, Synechococcus, Pleurocapsa, and Prochlorococcus), should be investigated further as potential TTX producers.
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