Infective Larvae Production and Development in Strongyloides and its role in the dauer hypothesis

The dauer hypothesis for the evolution of parasitism in nematodes, states that dauer larvae, a specialised third larval stage present in many free living nematodes, served as a pre adaptation to parasitism and evolved into infective larvae (L3i). Unique biological features like alternating free-living sexual and parasitic parthenogenetic generations make Strongyloides spp., which includes the human pathogen S. stercoralis, an attractive model to study parasitism and the evolution thereof. One of the most striking features is that all L3is are female, although they can be produced from sexual reproduction. The main aims of this thesis were to test the dauer hypothesis for the specific example of Strongyloididae and to understand how female only progeny are formed from sexual reproduction. While Strongyloides ratti is an attractive model species as it can be maintained within its natural host, its free living stages can only be extracted from fecal cultures and many tools that are standard in model organisms are not yet available, making experimentation difficult. Therefore, I firstly set out to increase the toolkit within S. ratti. I strongly improved the protocol to culture S. ratti on bacteria seeded agar plates, which led to highly increased fecundity and survival, allowing easier experimentation. Further I devised the first working procedures for RNAi in any species of Strongyloides. As S. ratti appears refractive to microinjection, I developed an RNAi by soaking protocol which works across multiple life stages and has minimal off-target effects. Using the aforementioned RNAi technique, I studied daf-12 natively within S. ratti (the first study of its kind within Strongyloides). By suppressing DAF-12 I found that many functions of this nuclear hormone receptor are conserved between C. elegans and S. ratti. In particular, daf-12(RNAi) inhibited the formation of infective larvae as it does for dauer larvae in C. elegans. This demonstrates that the molecular mechanisms underlying dauer formation and L3i formation are conserved between species as phylogenetically distant as C. elegans and S. ratti , suggesting that DAF-12 it is a promising target for novel antihelminthics and providing support for the dauer hypothesis. In order to study the transition from free-living to parasitic, we need well-studied closely related free-living species to parasites of interest, of which we are currently lacking. After noticing that Rhabditophanes KR3021 (the closest known free living species to Strongyloides) produces dauer larvae and arrested J2 larvae (both previously unreported) under starvation conditions, I aimed to characterise this species and provide clearly lacking basic information. In addition to providing a detailed description of life cycle and morphology, I showed that the daf-12 pathway is also involved in dauer (but not arrested J2) formation making Rhabditophanes/Strongyloides a highly attractive system for the further evaluation of the dauer hypothesis. Next, by characterising spermatogenesis in S. ratti, S. papillosus and P. trichosuri (which does make males), I found that there is an uneven division resulting in viable X-bearing and non-viable nullo-X sperm. I showed that spermatogenesis itself is highly conserved within the Strongyloididae, from the presence of giant nuclei in the distal gonad, to expression and production of major sperm protein

The Effects of Different Substrates on the Quality of Seedlings and Yield in Organic Production of Lettuce

An experiment was conducted under glasshouse condition. Lettuce were sown in 5 different substrate: 100% earthworm; 60% earthworm + 40% potting soil; 85% earthwarm + 15% potting soil; 100% potting soil; certified organic substrate from the market (mix of white and black peat). The quality parameters of seedlings were noted: number of days from sowing to emergence, development of first leaf, total number of emerged plants. During vegetation are recorded dates about dynamics of growth. On the harvested yield were counted number of marketable and damaged leafs, diameter and height of head were determined and total weight of heads. Recorded results show that best substrate for production of seedlings of organic lettuce are mix of 60% earthwarm + 40% potting soil and commercial mix of white and black peats

Phytochemical and biological activity studies on Nasturtium officinale (watercress) microshoot cultures grown in RITA®^{®} temporary immersion systems

The main compounds in both extracts were gluconasturtiin, 4-methoxyglucobrassicin and rutoside, the amounts of which were, respectively, determined as 182.93, 58.86 and 23.24 mg/100 g dry weight (DW) in biomass extracts and 640.94, 23.47 and 7.20 mg/100 g DW in plant herb extracts. The antioxidant potential of all the studied extracts evaluated using CUPRAC (CUPric Reducing Antioxidant Activity), FRAP (Ferric Reducing Ability of Plasma), and DPPH (1,1-diphenyl-2-picrylhydrazyl) assays was comparable. The anti-inflammatory activity of the extracts was tested based on the inhibition of 15-lipoxygenase, cyclooxygenase-1, cyclooxygenase-2 (COX-2), and phospholipase A2. The results demonstrate significantly higher inhibition of COX-2 for in vitro cultured biomass compared with the herb extracts (75.4 and 41.1%, respectively). Moreover, all the studied extracts showed almost similar antibacterial and antifungal potential. Based on these findings, and due to the fact that the growth of in vitro microshoots is independent of environmental conditions and unaffected by environmental pollution, we propose that biomass that can be rapidly grown in RITA® bioreactors can serve as an alternative source of bioactive compounds with valuable biological properties

Metformin Prevents Nigrostriatal Dopamine Degeneration Independent of AMPK Activation in Dopamine Neurons

Metformin is a widely prescribed drug used to treat type-2 diabetes, although recent studies show it has wide ranging effects to treat other diseases. Animal and retrospective human studies indicate that Metformin treatment is neuroprotective in Parkinson’s Disease (PD), although the neuroprotective mechanism is unknown, numerous studies suggest the beneficial effects on glucose homeostasis may be through AMPK activation. In this study we tested whether or not AMPK activation in dopamine neurons was required for the neuroprotective effects of Metformin in PD. We generated transgenic mice in which AMPK activity in dopamine neurons was ablated by removing AMPK beta 1 and beta 2 subunits from dopamine transporter expressing neurons. These AMPK WT and KO mice were then chronically exposed to Metformin in the drinking water then exposed to MPTP, the mouse model of PD. Chronic Metformin treatment significantly attenuated the MPTP-induced loss of Tyrosine Hydroxylase (TH) neuronal number and volume and TH protein concentration in the nigrostriatal pathway. Additionally, Metformin treatment prevented the MPTP-induced elevation of the DOPAC:DA ratio regardless of genotype. Metformin also prevented MPTP induced gliosis in the Substantia Nigra. These neuroprotective actions were independent of genotype and occurred in both AMPK WT and AMPK KO mice. Overall, our studies suggest that Metformin’s neuroprotective effects are not due to AMPK activation in dopaminergic neurons and that more research is required to determine how metformin acts to restrict the development of PD

Mutations in VPS13D lead to a new recessive ataxia with spasticity and mitochondrial defects

Peer Reviewedhttps://deepblue.lib.umich.edu/bitstream/2027.42/145306/1/ana25220_am.pdfhttps://deepblue.lib.umich.edu/bitstream/2027.42/145306/2/ana25220.pd

Autophagy Induction as a Therapeutic Strategy for Neurodegenerative Diseases.

Autophagy is a major, conserved cellular pathway by which cells deliver cytoplasmic contents to lysosomes for degradation. Genetic studies have revealed extensive links between autophagy and neurodegenerative disease, and disruptions to autophagy may contribute to pathology in some cases. Autophagy degrades many of the toxic, aggregate-prone proteins responsible for such diseases, including mutant huntingtin (mHTT), alpha-synuclein (α-syn), tau, and others, raising the possibility that autophagy upregulation may help to reduce levels of toxic protein species, and thereby alleviate disease. This review examines autophagy induction as a potential therapy in several neurodegenerative diseases-Alzheimer's disease, Parkinson's disease, polyglutamine diseases, and amyotrophic lateral sclerosis (ALS). Evidence in cells and in vivo demonstrates promising results in many disease models, in which autophagy upregulation is able to reduce the levels of toxic proteins, ameliorate signs of disease, and delay disease progression. However, the effective therapeutic use of autophagy induction requires detailed knowledge of how the disease affects the autophagy-lysosome pathway, as activating autophagy when the pathway cannot go to completion (e.g., when lysosomal degradation is impaired) may instead exacerbate disease in some cases. Investigating the interactions between autophagy and disease pathogenesis is thus a critical area for further research

Modulating mitophagy in mitochondrial disease

Mitochondrial diseases may result from mutations in the maternally-inherited mitochondrial DNA (mtDNA) or from mutations in nuclear genes encoding mitochondrial proteins. Their bi-genomic nature makes mitochondrial diseases a very heterogeneous group of disorders that can present at any age and can affect any type of tissue. The autophagic-lysosomal degradation pathway plays an important role in clearing dysfunctional and redundant mitochondria through a specific quality control mechanism termed mitophagy. Mitochondria could be targeted for autophagic degradation for a variety of reasons including basal turnover for recycling, starvation induced degradation, and degradation due to damage. While the core autophagic machinery is highly conserved and common to most pathways, the signaling pathways leading to the selective degradation of damaged mitochondria are still not completely understood. Type 1 mitophagy due to nutrient starvation is dependent on PI3K (phosphoinositide 3-kinase) for autophagosome formation but independent of mitophagy proteins, PINK1 (PTEN-induced putative kinase 1) and Parkin. Whereas type 2 mitophagy that occurs due to damage is dependent on PINK1 and Parkin but does not require PI3K. Autophagy and mitophagy play an important role in human disease and hence could serve as therapeutic targets for the treatment of mitochondrial as well as neurodegenerative disorders. Therefore, we reviewed drugs that are known modulators of autophagy (AICAR and metformin) and may effect this by activating the AMP-activated protein kinase signaling pathways. Furthermore, we reviewed data available on supplements, such as Coenzyme Q and the quinone idebenone, that we assert rescue increased mitophagy in mitochondrial disease by benefiting mitochondrial function

Transposition of temporality in French, English and BCMS : between tense and aspect of the verb

L’objectif de la présente thèse est de mettre en évidence les différentes valeurs des temps verbaux du passé dans un texte littéraire, ainsi que les possibilités de leur traduction d’une langue à l’autre. L’étude est élaborée dans une perspective comparative, portant à la fois sur une langue romane (le français), une langue germanique (l’anglais) et une langue slave (les langues BCMS, à savoir bosniaque, croate, monténégrin et serbe). À partir d’un corpus de traductions de nouvelles rédigées en français (Boule de suif de G. de Maupassant), en anglais (The Dead de J. Joyce) et en BCMS (Enciklopedija mrtvih de D. Kiš), nous étudions l’emploi et le fonctionnement des temps du passé dans les textes narratifs, ainsi que leurs équivalents traduits dans les autres langues. L’analyse de travaux d’étudiants de la Faculté de philologie de l’Université du Monténégro nous permet ensuite de présenter les problèmes auxquels les traducteurs sont confrontés en transposant les temps du passé d’une langue à l’autre, aussi bien que les choix et les stratégies de contournement des obstacles rencontrés. Nous soulignons enfin l’importance de la formation des traducteurs non seulement dans les domaines des théories aspecto-temporelles de trois systèmes langagiers, mais aussi au niveau de divers modes discursifs, qui parfois déterminent, conditionnent et limitent l’emploi des temps du passé.The objective of this thesis is to highlight the different values of past tenses in a literary text, as well as the possibilities of their translation from one language to another. The study is developed from a comparative perspective, covering a Romance language (French), a Germanic language (English) and a Slavic language (BCMS, i.e., Bosnian, Croatian, Montenegrin and Serbian). Based on a corpus of translations of short stories written in French (Boule de suif by G. de Maupassant), in English (The Dead by J. Joyce) and in BCMS (Enciklopedija mrtvih by D. Kiš), we study the use and functions of past tenses in narrative texts, as well as their translation equivalents in other languages. Furthermore, the analysis of the works of students of the Faculty of Philology at the University of Montenegro allows us to present the problems that translators face when translating past tenses from one language to another, as well as the choices and strategies for overcoming the obstacles encountered. Finally, we underline the importance of educating translators not only in the area of aspecto-temporal theories of three language systems, but also with regard to the various discursive modes, which sometimes determine, condition and limit the use of past tenses

La transposition de la temporalité en français, anglais et BCMS : entre temps et aspect du verbe

The objective of this thesis is to highlight the different values of past tenses in a literary text, as well as the possibilities of their translation from one language to another. The study is developed from a comparative perspective, covering a Romance language (French), a Germanic language (English) and a Slavic language (BCMS, i.e., Bosnian, Croatian, Montenegrin and Serbian). Based on a corpus of translations of short stories written in French (Boule de suif by G. de Maupassant), in English (The Dead by J. Joyce) and in BCMS (Enciklopedija mrtvih by D. Kiš), we study the use and functions of past tenses in narrative texts, as well as their translation equivalents in other languages. Furthermore, the analysis of the works of students of the Faculty of Philology at the University of Montenegro allows us to present the problems that translators face when translating past tenses from one language to another, as well as the choices and strategies for overcoming the obstacles encountered. Finally, we underline the importance of educating translators not only in the area of aspecto-temporal theories of three language systems, but also with regard to the various discursive modes, which sometimes determine, condition and limit the use of past tenses.L’objectif de la présente thèse est de mettre en évidence les différentes valeurs des temps verbaux du passé dans un texte littéraire, ainsi que les possibilités de leur traduction d’une langue à l’autre. L’étude est élaborée dans une perspective comparative, portant à la fois sur une langue romane (le français), une langue germanique (l’anglais) et une langue slave (les langues BCMS, à savoir bosniaque, croate, monténégrin et serbe). À partir d’un corpus de traductions de nouvelles rédigées en français (Boule de suif de G. de Maupassant), en anglais (The Dead de J. Joyce) et en BCMS (Enciklopedija mrtvih de D. Kiš), nous étudions l’emploi et le fonctionnement des temps du passé dans les textes narratifs, ainsi que leurs équivalents traduits dans les autres langues. L’analyse de travaux d’étudiants de la Faculté de philologie de l’Université du Monténégro nous permet ensuite de présenter les problèmes auxquels les traducteurs sont confrontés en transposant les temps du passé d’une langue à l’autre, aussi bien que les choix et les stratégies de contournement des obstacles rencontrés. Nous soulignons enfin l’importance de la formation des traducteurs non seulement dans les domaines des théories aspecto-temporelles de trois systèmes langagiers, mais aussi au niveau de divers modes discursifs, qui parfois déterminent, conditionnent et limitent l’emploi des temps du passé