Microbial Ecology of Phototrophs in Iron-rich Boreal Shield Lakes

The early Earth oceans of the Archaean Eon (approx. 3.8-2.5 billion years ago) are thought to have been predominantly anoxic with ferruginous waters containing high levels of dissolved ferrous iron and low levels of sulfate. How primary production operated in these ferruginous waters, where early life would have evolved, remains poorly understood. Based on the global occurrence of massive banded iron formations that can be dated to as early as the Mesoarchaean Era, many speculate that biological processes helped to drive the oxidation of iron in the Earth’s oceans, which today are sulfate-rich and iron-poor. Along with oxygenic photosynthesis, where produced molecular oxygen can oxidize iron abiotically, alternative microbial processes such as photoferrotrophy, where ferrous iron serves as the direct electron donor for phototrophy, have been implicated in the deposition of banded iron formations. Other forms of anoxygenic phototrophy, such as phototrophic sulfide oxidation, could have also served as a means of primary production in early ferruginous waters. However, the diversity, ecology, and relative importance of anoxygenic forms of phototrophy in Archaean oceans remain poorly characterized given the current global prevalence of molecular oxygen. Boreal Shield lakes number in the millions across northern regions. Although most Boreal Shield lakes mix and re-oxygenate seasonally, an estimated 15% of these lakes develop anoxic bottom waters over the summer or winter months that gain high levels of dissolved iron while having low levels of sulfate. As such, these Boreal Shield lakes can potentially complement permanently anoxic and ferruginous lakes that have been used to gain insights about the properties of ferruginous waters but are rare globally. The research presented in this thesis aimed to characterize the diversity and ecology of phototroph communities in the ferruginous water columns of seasonally anoxic Boreal Shield lakes. By performing a multi-year lake survey at the International Institute for Sustainable Development Experimental Lakes Area (IISD-ELA) combining high-throughput DNA/RNA sequencing, enrichment cultivation, physicochemical characterization, and rate measurements, the goal of this research was to clarify the metabolic diversity of phototrophy, physicochemical controls on phototrophs, and the relative importance of different modes of phototrophy in Boreal Shield lake anoxic zones with implications for modern ecology and for study of early Earth biogeochemistry. Metagenomes were sequenced from preliminary DNA samples collected from Lakes 227 and 442 at the IISD-ELA to analyze the functional gene content of the lake microbial communities (Chapter 2). Metagenomes were assembled and contigs binned to recover high-completeness and low-contamination draft genome bins. The genetic potential for sulfide oxidation, indicated by the dsrA gene, was found in three of the four recovered and curated genome bins classified to the Chlorobia class, but the fourth genome bin was found to contain the cyc2 candidate marker gene for iron oxidation and was unique to Lake 227. Two new species of Chlorobia were enriched in sulfide-containing medium from Lakes 227 and 304. Recovered draft genome bins of both novel Chlorobia members contained both the dsrA and cyc2 genes, indicative of the potential for both ferrous iron and sulfide oxidation. The Lake 304 strain, provisionally named, “Candidatus Chlorobium canadense”, was subsequently incubated in ferrous iron-containing medium yet did not show signs of photoferrotrophic activity. Nevertheless, robust methods for the detection of Chlorobia-associated cyc2 homologs were developed, and analysis of reference genomes indicated that the presence of the cyc2 gene is associated with photoferrotrophic potential. Genomic potential for iron- and sulfur-cycling processes were also detected among several other recovered genome bins outside the Chlorobia. Overall, evidence for sulfide-oxidizing phototrophy was found in the lakes despite low sulfate levels, along with genomic potential for photoferrotrophy, and “Ca. Chl. canadense” was enriched to allow for future study of the functional role of cyc2 among the Chlorobia. Expanding on initial results from cultivation and metagenome analyses, nine Boreal Shield lakes at the IISD-ELA with diverse physicochemical characteristics were surveyed in the summer and/or fall from 2016-2018 to understand the broader distribution of phototrophs among ferruginous lake systems (Chapter 3). Lake anoxic zones samples generally grouped into two categories based on microbial community composition as inferred from 16S rRNA gene and metagenome data. One of those categories was associated with high relative abundances and diversities of anoxygenic phototrophs within the Chlorobia and Chloroflexota phyla and included three lakes. Lake categories based on microbial community composition generally matched categories determined via hierarchical clustering based on lake physicochemical parameters. Compared to the other surveyed lakes, the three detected “phototroph-rich” lakes tended to have low surface area:depth ratios with high dissolved organic carbon content, high dissolved iron content, and low levels of sulfate in their anoxic waters. Anoxic water column metagenomes from the three lakes contained high relative abundances of Chlorobia members along with Chlorobia-associated cyc2 and dsrA genes. Metatranscriptome sequencing revealed that Chlorobia members comprised nearly 50% of total gene expression in the upper anoxic zones of these three lakes based on the rpoB single-copy taxonomic marker gene. Among Chlorobia members, the dsrA gene for phototrophic sulfide oxidation was highly expressed, whereas the cyc2 gene was expressed at lower levels for two lakes (Lakes 221 and 304) and minimally expressed for the third (Lake 227). Rate incubation data from Lake 227 suggested that oxygenic phototrophy was the dominant contributor to iron oxidation even 1 m below the measured oxic/anoxic zone boundary. Measured iron oxidation rates exceeded measured rates of iron reduction at all measured regions of the upper anoxic water column. Sulfide-oxidizing phototrophy was also detected and was tightly coupled to sulfate reduction. Photoferrotrophy was not detected but may have occurred at rates less than the observed iron reduction rate. Metatranscriptome data also revealed high expression of the particulate methane monooxygenase (pmoA) gene deep in the anoxic zone of Lake 227, implying that novel forms of methane oxidation occur in this lake. Altogether, the data demonstrate that Boreal Shield lakes commonly contain high relative abundance and diverse phototroph communities in their seasonal anoxic zones. In addition, the lake anoxic zones appear to host active and cryptic sulfur cycles, along with the potential for iron oxidation/reduction and methane cycling. Enrichment cultivation in a ferrous iron-containing medium from Lake 227 allowed for the recovery and characterization of a novel anoxygenic phototrophic Chloroflexota member, provisionally named “Candidatus Chlorohelix allophototropha” (Chapter 4). This organism performs phototrophy using a distinct fourth clade of Type I photosynthetic reaction center (RCI) protein, despite placing sister taxonomically to Type II reaction center (RCII)-utilizing Chloroflexota members. “Ca. Chx. allophototropha” contains chlorosomes, uses bacteriochlorophyll c, and encodes the FMO protein like other RCI-utilizing phototrophs in the Chlorobiales and Chloracidobacterales orders. “Ca. Chx. allophototropha” also encodes the potential for carbon fixation using the Calvin-Benson-Bassham (CBB) cycle, unlike all known RCI-utilizing phototrophs. The discovery of “Ca. Chx. allophototropha”, as the first representative of a novel Chloroflexota order (i.e., “Ca. Chloroheliales”), sheds light on longstanding questions about the evolution of photosynthesis, including the origin of chlorosomes among RCII-utilizing Chloroflexota members. The Chloroflexota is now the only phylum outside the Cyanobacteria containing genomic potential for both major classes of photosynthetic reaction center and can thus serve as an additional system for exploring fundamental questions about the evolution of photosynthesis

n-3系脂肪散を強化した食事療法が有効と考えられた肺気腫の一例

An effective treatment for the advanced stages of chronic obstructive pulmonary disease (COPD) has not been established yet. We report our recent experience of one patient with pulmonary emphysema treated by dietary supplementation of n-3 fatty acid for two months. He presented improvements in clinical symptoms and pulmonary function, and suppression of leukotriene B(4) generation by peripheral leukocytes. We consequently suppose that dietary treatment with n-3 fatty acids (perilla seed oil) may offer benefits for the treatment of pulmonary emphysema by competitively inhiabiting the conversion of arachidonicacid to leukotrienes and prostanoids.今回我々は,肺気腫の症例に対してn-3系脂肪酸を強化した食事療法をおこない,臨床症状,呼吸機能検査所見ともに速やかに改善を認め,同時に白血球のロイコトリエンB4産生能が著明に減少した一例を経験したので報告する｡ 症例は67歳,男性｡主訴は労作時呼吸困難｡【第一回目入院】3カ月間入院し,薬物療法,温泉を用いた理学療法を行った｡自覚症状はやや改善が見られたが,呼吸機能検査所見の改善は得られなかった｡【第二回目入院】1年後に再入院｡n-3系脂肪酸強化食事療法も併用した｡自覚症状および,呼吸機能検査上,FVC,FEV1.0,PEFなどに改善を認めた｡n-3系脂肪酸はアラキドン酸代謝を通してロイコトリエン合成に関与すると推定されるが,経渦中に白血球のLTB4産生能の減少を認めた｡この症例は肺気腫に対するn-3系脂肪酸強化食 事療法の有用性が示唆され,病態を考える上でも興味深いと考えられた

Noncontact Impedance Control for Redundant Manipulators

This paper proposes an impedance control methodthat can regulate a virtual impedance between a robot manipulatorand external objects using visual information. Theconventional impedance control method is not useful in somecases where no interaction force between the arm and its environmentexists, although it is one of the most effective controlmethods for manipulators in contact with the environment. Usingthe proposed method, we can control the manipulator motionbased on the virtual impedance before contact with the objects.The validity of the proposed method is verified through computersimulations and experiments using a direct-drive robot

Relationship between Eimeria tenella associated-early clinical signs and molecular changes in the intestinal barrier function

The major clinical signs of coccidiosis in chickens due to Eimeria parasite are diarrhea and bloody feces. Previous studies showed that the impairment of the intestinal epithelial barrier and the elevation of the intestinal permeability are causes of clinical signs associated with coccidia challenges. Nevertheless, the information about molecular changes of the epithelial barrier at the early stage of the infection with a specific Eimeria species has not been mentioned. Hence, this study aims to elucidate the temporal relationships between epithelial barrier conditions and clinical signs in chickens infected with Eimeria tenella over the time from the earliest stages of infection. White Leghorn chickens were inoculated with 1 × 104 oocysts of E. tenella. Thereafter the chickens were monitored for their daily clinical signs through observation, and between 5 dpi to 10 dpi, feces were collected for oocysts counting. Chickens were then administrated with fluorescein isothiocyanate-dextran (FITC-d) for gastrointestinal permeability test and tissues were collected each day for histopathological observation and total RNA extraction. Finally, the mRNA expression levels of the tight and adherens junction genes and cytokine genes were evaluated using the quantitative real-time polymerase chain reaction (qRT-PCR). In this study, clinical signs such as diarrhea and bloody feces were observed concurrently from 3 to 8 dpi. Histopathology changes such as severe inflammation, hemorrhage, and epithelial desquamation were identified in the cecum specimens. The FITC-d level in the E. tenella-infected group was significantly higher than in the control group. In the infected group, the expression of claudin-2 gene was also upregulated, whereas the expressions of claudin-3 and E-cadherin genes were decreased as compared to the control group. These results implied that clinical signs of avian coccidiosis were associated with the intestinal barrier disruption via changes in expression levels of claudins and E-cadherin at the intestine

サーモグラフィーによる体表面温度の測定 3.体表面温度の回復率と末消血流量との相関

The body surface peripheral circulation in 12 cases,　including 9 patients with　diabetes mellitus who were suffering coldness, numbness or pain in their feet, and 3 healthy volunteers was examined using Laser-Doppler blood flowmetry. At the same time, the body surface temperature was estimated by thermography. Thermographic results were analyzed quantitatively by calculating a recovery ratio as : Recovery ratio = [Total counts of thermography(Pixels) over temperature (T) after cold loading] ÷ [Initial counts over T before cold loading] x 100(%). The recovery ratio and the blood flow were correlated, r=0.68, p<O.01. The peripheral circulation of 16 patients with diabetes mellitus was observed at three different conditions including, l)placed at room temperature at 20℃for 15 min, 2) submerged and warmed for 5 min in a hot bath at 36℃(i.e. hot loading), and 3) submerged and cooled for 5 min in a water bath at 20℃ (i.e. cold loading). Three different baseline temperatures, 26℃, 27℃ and 28 ℃, were used in processing the　thermographic results into pictures. The highest correlation (r=0.59, p=0.0002) was obtained under the condition of cold loading using a baseline temperature limitation of27℃.The difference ratio (%) of blood flow was calculated as the blood flow at cold loading divided by the blood flow at hot loading in these 16 patients. The difference　ratio of the blood flow and the recovery ratio of thermography were correlated, r=0.46, p<O.OOO1. We found a strong correlation between the results of Laser-Doppler blood flowmetry and one of thermographic methods used to monitor peripheral circulation in patients with diabetes mellitus. Cold loading using a baseline temperature limitation of 27℃ were recommended for further examinations. Patients with low blood flow as well as　with large differences in their peripheral circulation between cold loading and hot　loading had severe coldness in their body surface temperature. We showed the usefulness of the results of thermography, when quantified by picture processing using computer software, in relation with the results of Laser-Doppler blood flowmetry.末梢神経障害を有する糖尿病患者の末梢循環障害の程度を数量的に検討する目的で､下肢に冷感ならびにしびれ感または疼痛を訴える糖尿病患者9症例と健常ボランティア3例の計12例(平均年齢59歳)についてサーモグラフィーを用いて体表面温度を測定した｡更に,サーモグラフィーで得られた結果と末梢皮膚血流量をレーザードプラー血流計を用いて測定して得られた結果と比較した｡サーモグラフィーによる測定で得られた結果は回復率として数量化して表示された｡回復率の算出方法は回復率= [冷水負荷後の特定温度27℃以 上の体表面温度のサーモグラフィーのPixelの総数]÷ [冷水負荷前の特定温度27℃以上の体表面温度のサーモグラフィーのPixelの総数]×100%で求めた｡レーザードプラ-血流計を用いて測定して得られた末梢血流量は左右それぞれ5カ所,計10カ所の測定値の平均で表示した｡その結果,末梢皮膚温度の回復率と末梢皮膚の血流量との問には正の相関関係(r=0.68,p<0.01)が認められた｡次に,末梢皮膚血流量について,室温20℃安静15分後,温水36℃浸水負荷10分後,冷水20℃浸水負荷30分後の異なる3条件について､またサーモグラフィーで得られた結果を,画像処理の過程で用いられた,26℃,27℃,28℃の3つの異なる特定温度との関連について検討を行なった｡対象は,下肢に冷感ならびにしびれ感または疼痛を訴える糖尿病患者16症例(平均年齢69歳,平均HbAIC9.6%)について測定した｡その結果,末梢皮膚 血流量は冷水20℃浸水負荷30分後に測定して得られた結果と,回復率は特定温度27℃で画像処理して得られた結果とが最も相関が高い(r=0.59,p=0.0002)ことが示された｡`次に,相関が高い条件は,室温20℃安静15分後に血流量を測定した場合(r=0.483,p=0.0002)であった｡そして温 水36℃浸水負荷10分後に測定して得られた結果とが最も相関関係が低い結果となった｡更に,冷水20℃浸水負荷30分後に測定して得られた結果を温水36℃浸水負荷10分後に測定して得られた結果で割った比を%で表示したところ回復率とこの比との間には正の相関関係(r=0.46,p<0.0001)が認められた｡このことから,温水36℃負荷時と,冷水20℃負荷時との差が大きい患者 において末梢皮膚温度の低下が著しいことが示された｡これまで悲観血的に測定されてきたサーモグラフィーによる末梢循環の数量的評価の試みは,レーザー血流計による結果と組み合わせることで,両者の間に正の相関関係が示されたことにより,今後,数量化された客観的評価を可能にした