Developing whole-cell biosensors for microbiome engineering applications

It is becoming increasingly apparent that the microbiota has a profound effect on human health and disease. Modern synthetic biology provides tools that can be used to engineer new diagnostic and therapeutic circuits- facilitating microbiome engineering and the creation of engineered biotherapeutics. These engineered biotherapeutics have the potential to expand our knowledge of microbial communities, host-microbe interactions and human health. However, to achieve these ambitious goals several challenges remain to be solved. These involve the creation of novel model systems and design strategies that can be used to characterise and improve these engineered strains. The primary focus of this thesis are whole-cell biosensors, that can be used to monitor molecules relevant to human health. Within this work I develop a novel model system, based on the Caenorhabditis elegans nematode that can be used to characterise biosensor strains in vivo. Through the developed protocols I use the nematode model to show that ratiometric biosensors can detect and report on changes within the C. elegans digestive tract. This model could be used to improve engineered biosensor strains, while also expanding our understanding of nematode biology and host-microbe interactions. In addition, I engineer a range of new ratiometric plasmids that can be used in conjunction with the C. elegans model system in future. Finally, I develop a range of acetoacetate-inducible biosensors; while also exploring methods of rationally improving two-component system biosensors. Two component systems are a common sensing mechanism that can be used to create a range of biosensors, therefore methods of rationally improving these biosensors would be an invaluable tool. Overall, it is hoped that the tools developed within this thesis can be used to further engineer whole-cell biosensors, which may help expand our knowledge of host-microbe interactions and human health

Identification and characterization of effectors secreted from sedentary endoparasitic phytonematodes

Sedentary endoparasitic phytonematodes are a group of taxa comprised of cyst nematodes (Heterodera and Globodera spp.) and root-knot nematodes (Meloidogyne spp. ), which are some of the most economically important crop pathogens on earth. These pests infect plant roots by creating elaborate feeding sites around the vasculature, which diverts nutrients away from the plant to feed the nematode and causes yield reduction in the plant. The nematodes create their feeding sites by delivering effector proteins into plant tissues. Effectors interact with plant components to modify development, metabolism, and defense pathways within plant cells, ultimately forming and maintaining the feeding site within the host root. In order to find ways to mitigate the damages caused by these nematode pathogens, it is vital to identify nematode effectors and understand how effector proteins are able to manipulate the plant host. This dissertation first summarizes what is currently known about nematode effector proteins and then contributes to that body of knowledge. Our data show that two effectors from Heterodera schachtii (Hs4E02 and Hs25A01) are likely to function in the plant−nematode interaction by binding to plant proteins. Through the preparation and mining of gland transcripts we have also identified 18 additional putative effectors that are expressed specifically within the esophageal gland cells of Meloidogyne incognita during infective life stages. Two of these putative effectors are part of the major avirulence protein (MAP) family of effectors. Interestingly, we were able to identify conserved amino acid motifs within this effector family that resemble plant CLAVATA3/ESR (CLE) signaling peptides found in plants and cyst nematodes. These data indicate that the MAP effector family from root−knot nematodes may manipulate plant developmental signaling in a manner analogous to the CLE effectors secreted from cyst nematodes

Evaluation of Amino Acid Composition as a Geochronometer in Buried Soils on Mount Kenya, East Africa

A sequence of surface and buried paleosols from the slopes of Mount Kenya, East Africa, has been identified and dated by radiocarbon and amino acid dating techniques in order to elucidate the Quaternary history of the area. Buried paleosols vary in radiocarbon age from 900 to > 40,000 yrs BP. They have developed in glacial and periglacial deposits of variable texture, consisting of a high percentage of clasts of phonolite, basalt and syenite. All but two paleosols are located in the Afroalpine zone (above 3200 m). D/L ratios of amino acids in Ab horizons were determined in order to establish their reliability for relative age dating. Alanine, aspartic acid, glutamic acid, leucine, valine, and phenylalanine were routinely analyzed. Aspartic acid, as in other cases, proved reliable yielding remarkably consistent results, with higher ratios corresponding to increasing age. Other acids analyzed showed distinct trends, although not as convincing as aspartic acid. In most cases, the aspartic acid ratio/ age relationships were supported by radiocarbon dates. D/L ratios of aspartic acid varied from approximately 0.07 for modern samples, to approximately 0.45 in samples > 40,000 years old.On a identifié et daté, au radiocarbone et à l'aide de techniques de datations à l'acide aminé, des paléosols enfouis et de surface afin de comprendre l'évolution quaternaire de la région. Les datations au radiocarbone des paléosols enfouis varient de 900 à plus de 40 000 BP. Les sols se sont développés dans des dépôts glaciaires et périglaciaires de différentes textures, constitués de fragments de roches détritiques, de phonolite, de basalte et de syénite. Tous les paléosols, sauf deux, sont situés dans la zone afroalpine (au-dessus de 3200 m). On a déterminé par racémisation les rapports D/L des acides aminés dans les horizons Ab en vue d'évaluer leur fiabilité pour la datation des âges relatifs. On a fait l'analyse de l'alaline, de l'acide aspartique, de l'acide glutamique, de la leucine, de la valine et du phénylalaline. L'acide aspartique, comme dans d'autres cas, a donné des résultats particulièrement satisfaisants, les quotients plus élevés correspondant aux âges les plus anciens. D'autres acides montraient des tendances bien distinctes, mais moins convaincantes que dans le cas de l'acide aspartique. Ainsi, dans la plupart des cas, les relations quotients/âges de l'acide aspartique étaient corroborées par les datations au radiocarbone. Les rapports D/L de l'acide aspartique variaient d'environ 0,07 pour les échantillons modernes à environ 0,45 pour les échantillons de plus de 40 000 ans.Eine Série von an der Oberflâche Negenden und begrabenen Palàobôden von den Hàngen des Mount Kenya, Ost-Afrika, wurde mittels Radiokarbon- und Aminosâuredatierungs-techniken identifiziert und datiert, um die Geschichte dieses Gebiets im Quaternàr zu erhellen. Das durch Radiokarbon bestimmte Alter der begrabenen Palàobôden variiert von 900 bis > 40,000 Jahren v.u.Z. Dièse Bôden haben sich in glazialen und periglazialen Ablagerungen verschiedener Beschaffenheit entwickelt, welche zu einem hohen Prozentsatz aus Trùmmern von Phonolith, Basait und Syenit bestehen. AuBer zweien befinden sich aile Palàobôden in der afroalpinen Zone (oberhalb 3200 m). Die D/L Anteile der Aminosâuren in den Ab-Horizonten wurden bestimmt, um ihre Verlâpiichkeit bei der relativen Altersbestimmung festzustellen. Alamin, aspartische Sàure, Glutamin-Sàure, Leuzin, Valin und Phenylalanin wurden laufend analysiert. Wie in anderen Fallen erwies sich die aspartische Sàure als verlàpiich, indem sie bemerkenswert bestàndige Ergebnisse ergab, bei denen die hôheren Quotienten dem hôheren Alter entsprachen. Andere analysierte Sàuren zeigten ausgepràgte Trends, wenn auch nicht so ùberzeugend wie die aspartische Sàure. In den meisten Fallen wurden die Beziehungen Quotient/Alter der aspartischen Sàure durch Radiokarbondatierungen gestùtzt. Die D/L Anteile der aspartischen Sàure variierten von ungefâhr 0.07 fur moderne Proben bis ungefâhr 0.45 in Proben, die > 40,000 Jahre ait sind

Whole-of-system approaches to physical activity;

Background: The literature on whole of system approaches (WSAs) has been largely theoretical in focus. The Australian Systems Approaches to Physical Activity is a national project designed to contribute a practical implementation focus to such approaches at the population level. Methods: National meetings were convened with federal and state government sector stakeholders to identify physical activity (PA) related policies and programs. Policies and programs were audited to develop an understanding of the existing PA system. A WSA conceptual map for PA was developed using feedback from system stakeholders, existing WSAs, and related work in obesity. Results: Completion of the policy audit has revealed key areas of need regarding policy governance, coordination, financing, and evaluation. An initial WSA conceptual map for Australia has been developed incorporating governance, translation, and advocacy. Stakeholder co-production of an integrated framework for PA and design plans for a community of practice knowledge hub has commenced. Conclusions: In Australia, Australian Systems Approaches to Physical Activity project partners have developed a conceptual whole of systems map that is guiding progress beyond the theoretical to application in the real world: A national PA policy audit, co-production of an integrated PA policy framework, and planning for a PA community of practice knowledge hub.</p

Edith Abbott Was Right: Designing Fieldwork Experiences for Contemporary Health Care Practice

Successful social work practice in the contemporary, economically driven, health care environment demands unprecedented levels of technical competence, initiative, creativity and conceptual sophistication. Fieldwork plays a critical role in social work education for such demanding practice by providing interns initial opportunities to apply their newly acquired knowledge, skills and abilities. This article discusses the contribution of fieldwork to the preparation of social work practitioners and presents two programs that may serve as alternative models of fieldwork. Observations of the impact of these models, including a summary of two evaluations of one model are presented. We contend that a fieldwork structure using rotations may increase the value of fieldwork for students both academically and in the employment process. Correct citation for final version of manuscript is: Spitzer, W., Holden, G., Cuzzi, L. C., Rutter, S., Chernack, P., & Rosenberg, G. (2001). Edith Abbott was right: Designing fieldwork experiences for contemporary health care practice. Journal of Social Work Education, 37, 79-90

Members of the Meloidogyne Avirulence Protein Family Contain Multiple Plant Ligand-Like Motifs

Sedentary plant-parasitic nematodes engage in complex interactions with their host plants by secreting effector proteins. Some effectors of both root-knot nematodes (Meloidogynespp.) and cyst nematodes (Heterodera and Globodera spp.) mimic plant ligand proteins. Most prominently, cyst nematodes secrete effectors that mimic plant CLAVATA3/ESR-related (CLE) ligand proteins. However, only cyst nematodes have been shown to secrete such effectors and to utilize CLE ligand mimicry in their interactions with host plants. Here, we document the presence of ligand-like motifs in bona fide root-knot nematode effectors that are most similar to CLE peptides from plants and cyst nematodes. We have identified multiple tandem CLE-like motifs conserved within the previously identified Meloidogyneavirulence protein (MAP) family that are secreted from root-knot nematodes and have been shown to function in planta. By searching all 12 MAP family members from multiple Meloidogyne spp., we identified 43 repetitive CLE-like motifs composing 14 unique variants. At least one CLE-like motif was conserved in each MAP family member. Furthermore, we documented the presence of other conserved sequences that resemble the variable domains described in Heterodera and Globodera CLE effectors. These findings document that root-knot nematodes appear to use CLE ligand mimicry and point toward a common host node targeted by two evolutionarily diverse groups of nematodes. As a consequence, it is likely that CLE signaling pathways are important in other phytonematode pathosystems as well

The Journal of the Friends' Historical Society vol. 5 No. 1

1. Notices. 2. Notes and Queries. 3. The Westmorland and Swaledale Seekers in 1651. 4. Extracts from the Minute Book of the Sufferings of Friends in Mansfield. 5. Reminiscences of the Friends' Meeting, Manchester. 6. Women Ministers stopped by Highwaymen. 7. Presentations of Quakers in Episcopal Visitations, 1662-1679. 8. Elisha Bates. 9. Keye-Worsley Marriage Certificate, 1666. 10. Thomas Areskine, Brewer, of Edinburgh. 11. Meeting Records. 12. A Glimpse of Ancient Friends in Dorset I. 13. Distribution of Literature in Cornwall, 1734. 14. William White, M.D. F.R.S. of York. 15. Friends in Barbadoes. 16. Some Quaker Teachers in 1736. 17. Friends in Current Literature. 18. Editors' Note. 19. Anecdote of Obed Cook, Schoolmaster. 20. Early Quaker Booksellers of York