Genetic variability of proteins involved in metronidazole metabolism and detoxification in Giardia lamblia

Metronidazole (MTZ) treatment-refractory Giardia lamblia infections is an emerging global problem. Recent studies show that prevalence rates of refractory infections are 10-20 % and can be as high as 46 %. The recent global increase of treatment refractory Giardia infections, suggest an emergence of strains with certain genetic inheritable traits. Up until now, no specific genetic marker of MTZ-resistance has been discovered in Giardia and most in vitro studies have utilized laboratory-derived sub-assemblage AI isolates, which rarely infects humans. The overall aim of this thesis was to investigate and analyze single nucleotide variants (SNVs) in proteins that are responsible for activation or inactivation of MTZ, and involved in the detoxification of reactive oxidative species and reactive nitrosative species in clinical isolates of Giardia. In this thesis we address the presence of genetic variation, especially amino acid altering, non-synonymous(ns) SNVs, of 29 genes. The variation was identified by Illumina whole genome sequencing of 20 recent, clinical, cultured sub-assemblage AII and assemblage B isolates, while cloning and Sanger sequencing was performed to identify alleles of the three genes flavohemoprotein (gFlHb), nitroreductase (NR) 1 and 2, the two latter were explored in eight non-cultured clinical isolates as well. Further, gFlHb was investigated for its potential variable copy number by mining PacBio de novo assembled genomes in eight isolates. Our findings disclose high frequencies of genetic variations in many of the 29 genes, where nsSNVs in assemblage B were especially high in ferredoxins (fds), NR1 and NR2, thioredoxin peroxidase and NADH oxidase. gFlHb was found to be a variable copy number gene (2-6 copies/isolate) with high genetic variation. Moreover, we identified truncated NR1 alleles in one MTZ refractory isolate, while two susceptible isolates, one homozygotic, had truncated NR2 alleles. The considerable genetic variation, coupled with findings of variable copies of gFlHb and dysfunctional NR proteins in recent clinical isolates of Giardia show a potential for identification of genetic markers for MTZ susceptibility/resistance. Future studies will help to decipher the effects of the mutations and could include functional assays and obtaining samples from a larger, more well-defined patient cohort.Doktorgradsavhandlin

Kryosfæriske massevariasjoner fra GRACE

The Earth’s gravity field is constantly changing due to mass redistribution from ice melting in the cryosphere and geophysical processes. With use of data from dedicated gravity satellite missions variations in the Earth’s gravity field over a time series can be determined. This thesis tries to give an explanation on how the Earth’s gravity field changes on a global scale and regionally with the use of data from the GRACE-mission over a time period. With comparison between three different GRACE-solutions and the use of SLR-observations, the mass changes over Greenland has been estimated. In addition, a further look into the lower degree spherical harmonics has been studied, to explain and give a conclusion of how the lower degree spherical harmonics from GRACE are affecting the calculations of the gravity field. GRACE-data has its uncertainties in the lower degree spherical harmonics. The C 20 -coefficients from GRACE has large variances and therefore needs to be replaced by SLR-values when looking at global variations. Trend calculations shows how post glacial rebound(GIA) influence Canada and Fennoscandia, and ice melting over Greenland, Alaska and Antarctica. The investigation of lower degree spherical harmonic coefficients shows that it is not significant how the C 20 -coefficients are processed when estimating mass changes over the area of Greenland. Due to restricted spatial resolution of the gravity field models and spatial averag- ing, it is hard to get a real estimate of the mass changes over a specific area. This is called the leakage effect and has been studied over the area of Greenland. The leakage effect has been estimated and restored, and mass change over Greenland has been estimated to be between -1832.76 km^3 and -2087.72 km^3 , depending on the model used. Jordas tyngdefelt er i stadig endring på grunn av masseforflytning fra isavsmelting i kryosfæren og geofysiske prosesser. Ved bruk av data fra gravimetrisatelliter kan disse variasjonene i jordas tyngdefelt bestemmes over en tidsserie. Denne oppgaven prøver å gi en forklaring på hvordan jordas tyngdefelt forandres globalt og regionalt ved bruk av data fra GRACE over en tidsperiode. Ved å sammenligne tre ulike GRACE-løsninger og ved bruk av SLR-observasjoner, har masseendringer over Grønnland blitt estimert. I tillegg har det blitt sett nærmere på de laveregrads sfærisk harmoniske koeffisientene, for å gi en forklaring på hvordan disse koeffisientene fra GRACE påvirker beregninger av jordens tyngdefelt. GRACE-data har sin usikkerhet i de laveregrads sfærisk harmoniske koeffisienter. C20-koeffisientene fra GRACE har store varianser og må derfor bli byttet ut med SLR-verdier når man ser på globale variasjoner. Trendberegninger viser hvordan postglasial landheving(GIA) påvirker områdene Canada og Fennoskandia, og i tillegg issmelting over Grønnland, Alaska og Antarktis. Ved å se nærmere på de laveregradskoeffisientene kan det bli observert at det ikke er av stor betydning hvordan man behandler C20-koeffisienter når masseendringer over Grønnland blir estimert. På grunn av begrenset romlig oppløsning og romlig glatting av tyngdefeltsmodeller, er det vanskelig å gi et realistisk estimat av masseendringer over et område. Dette blir kalt "leakage"-effekt. "Leakage"-effekten har blitt estimert og gjennopprettet, og masseendringer over Grønnland har blitt beregnet til å være mellom -1832.76 km^3 og -2087.72 km^3, avhengig av hvilken modell som har blitt brukt.M-GEO

Developing a flow cytometric method to characterize human T cell responses against the enteric protozoan parasite Giardia lamblia

Giardia lamblia is an enteric protozoan parasite, which causes infection in humans worldwide. The impact of the infection varies from asymptomatic carriers to severe disease such as malabsorption syndrome. Evidence for acquired immunity against Giardia infection has been found in previous studies. CD4+ T cell responses have been detected in humans, but data regarding cytokine producing profiles of these T cells is limited. This study aimed to develop a flow cytometric method to investigate Giardia-specific CD4+ T cell responses in individuals with recent giardiasis. Early cytokine profiles in addition to later surface markers and proliferation were combined to explore Giardia-specific CD4+ T cell immune responses by flow cytometry. In the development of the flow cytometric assay, fluorochrome-conjugated antibodies were titrated and different clones tested, detector voltages on the flow cytometer were adjusted, CellTrace proliferation dye labeling method and concentration was optimized, spectral overlap was minimized, compensation matrices were acquired and different fixation and permeabilization reagents were tested. To explore the function of the assay, cultures of peripheral blood mononuclear cells (PBMCs) from a group of individuals with recent giardiasis were stimulated with Giardia assemblage A and B sonicated soluble proteins (SSA and SSB), and responses were compared to responses in PBMCs from a group of low risk healthy controls. Early cytokine profiles in addition to later surface markers and proliferation were compared between these groups to explore Giardia-specific CD4+ T cell immune responses by flow cytometry. The first assay (day one assay) investigated cytokine expression of TNA-α, IFN-γ, IL-17A, IL-10 and IL-4 in effector memory CD4+ T cells (CD197-CD45RA-) after 24 hours of stimulation with Giardia soluble proteins and controls antigens. The other assay (day six assay) investigated proliferation by CellTrace dye dilution and activation markers HLA- DR, CD45RO, CD25 and CD26 after six days of stimulation. The results were analyzed in FlowJo, and statistical analysis was done using SPSS. Cytokine responses were stronger in the Giardia exposed group when stimulated with SSA and SSB, but only IL-17A production was found to be significantly elevated in this group. Two participants with current, on-going, giardiasis had markedly elevated production of all cytokines, except IL-4, in response to SSA and SSB, but not to control antigens. In the day six assay CD45RO and HLA-DR positive cell percentages were found to be significantly different between the groups when stimulated with SSA. A positive correlation between SSA and SSB induced effector memory CD4+ T cell cytokine production, as well as proliferation responses was found, indicating considerable cross-reaction between these two assemblages. In conclusion, we find that the developed assay performed well and can be used to assess Giardia-specific immunity, but it has some shortcomings. Although the assay showed generally higher responses in the giardiasis exposed group for most of the outcomes, only IL-17A production, and HLADR+CD45RO+ activation turned out to be statistically significant. Future studies using purified recombinant Giardia proteins as antigens may..

Studies on the stimulation of Atlantic salmon macrophage-like cells with emphasis on respiratory burst

Reactive oxygen species (ROS) production in macrophage-like cells is induced as an antimicrobial defence against invading pathogens. In this present study, we have explored how different stimuli and metabolic inhibitors affects the level of respiratory burst in Atlantic salmon (Salmo salar L.) head kidney macrophage-like cells. Cells stimulated in vitro by bacterial lipopolysaccharide (LPS) and ß-glucan showed increased production of ROS compared to unstimulated cells. Both stimulation and co-stimulation by curdlan (ß-glucan) induced a higher production of ROS compared to stimulation and co-stimulation by LPS. Metabolic inhibitors (developed for mammals) co-incubated with the stimulants did not, in most cases, perturb the level of ROS generation in the salmon macrophage-like cells. The NAD+ content as well as the NAD+/NADH ratio increased in curdlan, and LPS + curdlan stimulated cells compared to control cells, which indicated increased metabolic activity in the stimulated cells. Supporting these findings, gene analysis using SYBR green real-time quantitative PCR showed that the genes Arignase-1 and IL-1ß were highly expressed in the stimulated cells

Brain Play Some psychological perspectives in flute performance training

Masteroppgave utøvende musikk, klassisk- Universitetet i Agder, 2015Both the field of psychology and that of neurology can provide important insights into the musical functions of the brain. This thesis aims to provide a pragmatic theory of how to incorporate aspects of these fields into music performance and education in a flute context. It is divided into two parts where Part I focuses on the theoretical methodologies that can be used for this purpose, including neurology, cognitive and social psychology, philosophy and how habits affect our lives. Part II combines the conclusions from Part I into a three-point ‘checklist’ that aims to provide a practical way of applying the theories to real-life practise, performance and educational settings

Extensive reading guided by a librarian: L2 English development in primary school

The aim of this study is to investigate the effects of extensive reading in English as a second language under the guidance of a trained librarian has on the language development of a class of sixth graders, with Norwegian as their first language. The study is based on Vygotsky’s (cop. 1978) theory of Proximal Development and Krashen’s (1981, and cop. 1982) theories of language acquisition. The researcher explores (i) the correlation between the number of books read and the pupils’ language development during the research period, (ii) the number of readings in context of an extensive reading project, (iii) the pupils’ choice of literature, and (iv) the impact of recommendations by a librarian. This is a mixed methods intervention study, where the pupils in the research class are given more time for self-selected reading in English than common in the English curriculum in Norwegian sixth grade. The following measures are included (i) the Cambridge Young Learners English tests (Cambridge English, cop. 2018; and Cambridge English, cop. 2014), (ii) a questionnaire for the pupils’ parents and guardians to retrieve language and social background variables, (iii) reading logs where the pupils register what they read during the research period, (iv) short stories written by the pupils, (v) observations of classroom interaction, and (vi) conversations with the pupils and their teachers. The pupils’ results in the Cambridge Young Learners Tests provides quantifiable results which will be used to measure their language development, this will be supported by the findings in the short stories written by the pupils during the research period. The pupils have recorded how many and which books they read in their reading logs. The researcher has kept lists over the books that were brought and presented to the class, and the impact of the recommendations by a librarian, that is the researcher, can be found by comparing the pupils’ reading logs with the researcher’s lists. The researcher’s predictions are that pupils who read more are more likely to improve in language proficiency. The number of books the pupils read will vary significantly and will be influenced both by their language competence and their earlier reading habits and reading experiences. The pupils will probably choose books of similar types and themes to those they would choose in Norwegian, and most will read at least some comic books. The books recommended by the librarian will probably be read by many of the pupils. In line with the findings from the reading programs described by Day and Bamford (1998), Krashen, Lee and Lao, (2018), Mason and Krashen (2004), Salameh (2017), and Axelsson (2000), the research for this master’s thesis shows an increased language proficiency among the pupils in a Norwegian sixth grade class who took part in this intervention study with increased focus on extensive reading in English as a second language. While there is a correlation between numbers of books read during the research period and increased language proficiency, the connection is neither linear nor unambiguous: The pupils who read the least during the research period, showed the least progress in language skills as measured by the pre- and post- research tests, while also one of the top scorers registered reading few books

The surveillance and control programme for bonamiosis and marteiliosis in European flat oysters, Ostrea edulis, and blue mussels, Mytilus sp. in Norway in 2019

This report describes the results from The surveillance and control programme for bonamiosis and marteiliosis and research activities on Marteilia sp. in 2019. The programme is carried out by the Institute of Marine Research according to a contract with the Norwegian Food Safety Authority. Samples were collected from two oyster farms, one wild oyster population, two mussel farms and four wild beds, two mussel farms and five wild mussel populations. Samples were collected in April/May and in October, in order to be able to detect Bonamia sp. and Marteilia sp. during the periods when the potential prevalence is at the highest. Samples from one of the mussel farms and one wild bed were collected after reports of elevated mortality. Except from these samples, no abnormal mortalities were observed during the surveillance. Bonamia ostreae / B. exitiosa were not detected. There have been several reports on mortality or “disappearance” of mussels along the Norwegian coast. The reason(s) for the mortalities have not been determined. However, the parasite Marteilia sp. was detected for the first time in mussels, Mytilus edulis, collected at Bømlo, western Norway in 2016 and Tysnes in 2019. This has been followed up with an extended survey in several research projects. The results show that Marteilia sp. infecting mussels in Norway, Sweden and England are different from Marteilia refringens infecting flat oysters. The name Marteilia pararefringens has been proposed, and there is strong evidence that Marteilia refringens and Marteilia pararefringens sp. nov. are distinct parasites of bivalves and have different European distributions. The work will be continued in 2020 – 2021, linked to research on the distribution of M. pararefringens in wild mussels. We propose a revision of the surveillance programme combined with the establishment of a new model for health control in mollusk farms, application for disease free status for Norwegian flat oysters and a categorization of zones.publishedVersio

Summarizing the screening for Bonamia ostreae in Norwegian populations of flat oysters, Ostrea edulis

Surveillance and studies performed by The Norwegian Institute of Marine Research and the Norwegian Veterinary Institute has never revealed notifiable diseases in Norwegian populations of European flat oysters, Ostrea edulis. In 2006, microcells resembling the oyster parasite Bonamia sp. were observed during histopathological examination of tissue specimens of flat oysters, Ostrea edulis from the Arendal area, southern Norway. The cells were however not interprated as B. ostreae and PCR-analysis of samples from this oyster population were negative. In 2008, the EU reference laboratory (EURL) received samples from the Norwegian Veterinary Institute and reported one Bonamia sp. in a haemocyte from one oyster, based on microscopy. By real-time PCR, positive results were obtained from two oysters in one triplicate sample. Sequencing of the PCR products gave 100% identity with B. ostreae. After this diagnose, both the Norwegian Veterinary Institute and The Institute of Marine Research have monitored the population. The results are briefly reported here. The observed microcells were found in most samples since the sampling at the site was initiated, always at a low prevalence and intensity. No inflammation, pathology or reductions of the oyster's condition have been associated with the observation. The population appears healthy, with a normal reproductive cycle pattern. Several year classes have been present throughout the study period. Since 2009, more than 3 000 oysters have been examined. Bonamia sp. has never been detected, and all PCR assays have been negative.publishedVersio

Revisjonsprosessen som eit samarbeidsprosjekt mellom lærar og elev: Ein kvalitativ studie om dialogen mellom lærarkommentarar og elevrevisjonar i eit skriveforløp

Korleis går samhandlinga mellom lærar og elevar føre seg i ein revisjonsprosess? Formålet med dette masterprosjektet er å få meir kunnskap om handlingane og reaksjonane til lærarar og elevar i eit prosess- og revisjonsfokusert skriveforløp. Casestudien opnar for å sjå tekstmaterialet i lys av konteksten, og å sjå nærare på korleis undervisningsformer, roller og tekstideal kan ha påverka tilbakemeldings- og revisjonspraksisen. Datamaterialet er henta inn i eit skriveprosjekt i ein klasse på mellomtrinnet der elevane over tre veker skreiv ei forteljing. Elevane fekk undervisning i sjangertrekk og verkemiddel, og blei instruert i korleis dei kunne å gi og ta i mot tilbakemeldingar som dei kunne nytte til revisjonsarbeid. Prosjektet blei avslutta med to rundar med skriftleg lærarrespons, i samsvar med innhald og samanheng, og rettskriving og andre formsaker. Elevtekstane og dei tilhøyrande tilbakemeldingane dannar empirien i prosjektet, og desse har blitt analysert for å finne ut kva som kjenneteiknar handlingane til partane og korleis dei samhandlar. Analysen av tilbakemeldingane viser at lærarpraksisen både inviterer elevane til dialog om tekst, og er styrande gjennom spesifikke forslag og lukka spørsmålsformuleringar. Tilbakemeldingane syner eit tekstideal med fokus på utførlege skildringar, noko som mellom anna viser seg i at lærarkommentarane ikkje omfattar oppfordringar om å slette tekst. Elevane gjer relativt få eigeninitierte endringar, og revisjonane er tett knytt opp til responsen. Det er stor skilnad i elevgruppa når det gjeld mengde og kompleksitet i revisjonane. Medan nokre elevar klarar å følgje opp kommentarar godt, utfører andre berre små endringar eller unngår revisjonar. Årsaker til dette kan vere manglande forståing for språkbruken i eller intensjonen med lærarkommentarane. Revisjonsprosessen er prega av at elevane har liten erfaring med revisjon, og at lærarane styrer revisjonshandlingane til elevane. Gjennom tilbakemeldingar forsøker lærarane å skape dialog gjennom å stille seg undrande til elevtekstane, men nokre stader kan fokuset på læringsprosess bli overskygga av ønsket om eit godt produkt.

Rock flour as soil improvement

Befolkninga i verden er stadig økende. God jord og gjødsel er nødvendig for å kunne høste gode avlinger. Bruk av stenmjøl som jordforbedringsmiddel har lang historie, og har vært gjenstand for forsøk gjennom lang tid. Virkningen vil imidlertid kunne variere mye med uttakssted, tekstur og bruksområde. Denne oppgava skrives ut fra erfaringer etter startåret av et 5-årig feltforsøk, der stenmjøl fra 2 ulike pukkverk i Østfold spres på sur sandjord i Tune i Østfold, og ut fra aktuell litteratur om temaet. Det første året var svært tørt, og det var derfor få signifikante resultat å se, sjøl om det er gjort et bredt spekter av analyser av jord og avling. Analyser av stenmjølet sammen med funn gjort i tidligere forskning indikerer imidlertid at det bør kunne forventes signifikante resultat i løpet av de 5 åra forsøket er ment å vare. Dette vil være i tråd med erfaringer som konkluderer med at stenmjøl neppe er noen rask vei til gode avlinger, men kan gi et varig lager av viktige mineraler i jorda som kan bidra over flere år, spesielt på myr og sandjord utsatt for næringsmangel. Stenmjøl er et lovlig driftsmiddel i økologisk landbruk, og derfor spesielt interessant innafor denne landbruksretningen, men kan òg være aktuelt i konvensjonelt landbruk. For pukkprodusentene er det ofte et overflødig biprodukt forbundet med lagerkostnader. Å ta i bruk denne ressursen på en fornuftig måte vil derfor være til gjensidig nytte.The world’s population is constantly increasing. Good soil and fertilizers are necessary to be able to harvest good yields. The use of different types of rock flour as soil improvement has a long history, and has been subject to trials through many years. The effects can still vary a lot with the origin of the material, the texture and the way of use. This master’s thesis is written from experiences after the first season of a 5-year field trial, where rock flour from 2 different quarries in Østfold is spread on acid sandy soil in Tune In Østfold, and from relevant literature on the subject. The first year was extremely dry, therefore few significant results could be seen, even though a wide range of analysis of soil and yields has been done. Analysis of the rock flour, together with findings in previous research, still indicate that significant results could be expected during the 5 years the trial is supposed to last. This will be in accord with experiences concluding that rock flour is no fast solution to obtain good yields, but can give a long-term supply of important minerals for several years, in particular on peat and sandy soils subject to nutrient deficiency. Rock flour is a legal input in organic farming, and therefore considered of special interest in this agricultural direction, but can also be of interest in conventional agriculture. In quarries, rock flour is often an excessive by-product associated with storage costs. Utilizing this resource in a reasonable way will therefore be of mutual interest.submittedVersionM-P