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Investigating prevalence and geographical distribution of Mycoplasma sp. in the gut of Atlantic salmon (Salmo salar L.)
Get PDFThe fish gut microbiota has gotten considerable attention in recent years, and the microbes harboring the intestine of fish are thought to grant the host various effects related to size, metabolism, feeding behavior, and immune response. A Mycoplasma species has been discovered as highly abundant in the salmon gut. However, the resident strain has not yet been isolated. Knowledge regarding its colonization and the impact it may have on the host is, therefore, limited. This study aimed to map the prevalence and geographical distribution of Mycoplasma in the salmon gut and discover its potential role as part of the gut microbiota.
Salmon gut content was sampled for both cultivation purposes and direct DNA analyses in this project. Samples were collected from two salmon farms in Norway, Skjervøy (n = 23) and Bømlo (n = 19), and one in Chile (n = 20). A selection of Bømlo samples (n = 10) was cultivated in enriched growth medium. The prevalence of Mycoplasma at different geographical sites was investigated by analyzing the bacterial composition in the Bømlo and Chile samples using 16S rRNA gene sequencing. Moreover, selected samples from Bømlo (n = 4), Skjervøy (n = 7), and Chile (n = 1) were further processed for whole-genome shotgun sequencing to obtain genomic information of the salmon-associated Mycoplasma.
Mycoplasma was found abundantly in Norwegian salmon but was not detected in Chilean salmon. Thus, in this study, we observed a geographical difference (p = 0.00023) in the mycoplasmas’ prevalence in the gut of farmed Atlantic salmon. The underlying reasons for the absence of Mycoplasma in Chilean salmon must be further investigated to explain our findings. Further, we found that the salmon-associated Mycoplasma’s DNA was most frequently classified as M. penetrans, which may suggest relatedness between these species. Whether the salmon Mycoplasma exhibits pathogenic or protective characteristics is not known. However, given the seemingly large prevalence of mycoplasmas in salmon, it is likely they exist in the gut microbiota as commensals. Further research is necessary to discover potential negative or positive impacts the salmon-associated Mycoplasma might have on the physiology and immunology of the fish.Tarmmikrobiotaen til fisk har fått auka merksemd dei siste åra, og mikroorganismane som utgjer denne har truleg innverknad på verten relatert til storleik, metabolisme, fôringsåtferd og immunrespons. Ein Mykoplasma-art har blitt oppdaga i rikelege mengder i laksetarmen. Arten er enno ikkje isolert, og det er lite kunnskap om denne bakterien si kolonisering, og om verknaden den kan ha på verten. Målet med denne studien var difor å kartlegge utbreiinga, samt den geografiske fordelinga av Mykoplasma i laksetarm, og å undersøke kva rolle denne bakterien potensielt har som del av tarmmikrobiotaen.submittedVersionM-BIOTE
Going Beyond Counting First Authors in Author Co-citation Analysis
Get PDFThe present study examines one of the fundamental aspects of author co-citation analysis (ACA) - the way co-citation
counts are defined. Co-citation counting provides the data on which all subsequent statistical analyses and mappings
are based, and we compare ACA results based on two different types of co-citation counting - the traditional type that
only counts the first one among a cited work's authors on the one hand and a non-traditional type that takes into
account the first 5 authors of a cited work on the other hand. Results indicate that the picture produced through this non-traditional author co-citation counting contains more coherent author groups and is therefore considerably clearer. However, this picture represents fewer specialties in the research field being studied than that produced through the traditional first-author co-citation counting when the same number of top-ranked authors is selected and analyzed. Reasons for these effects are discussed
Variations on the Author
Get PDF“Variations on the Author” discusses two of Eduardo Coutinho’s recent films (Um Dia na Vida, from 2010, and Últimas Conversas, posthumously released in 2015) and their contribution to the general question of documentary authorship. The director’s filmography is characterized by a consistent yet self-effacing form of authorial self-inscription: Coutinho often features as an interviewer that rather than express opinions propels discourses; an interviewer that is good at listening. This mode of self-inscription characterizes him as an author who is not expressive but who is nonetheless markedly present on the screen. In Um Dia na Vida, however, Coutinho is completely absent form the image, while Últimas Conversas, on the contrary, includes a confessional prologue that moves the director from the margins to the center of his films. This article examines the ways in which these works stand out in the filmography of a director who offers new insights into the notion of cinematic authorship
Appropriate Similarity Measures for Author Cocitation Analysis
Get PDFWe provide a number of new insights into the methodological discussion about author cocitation analysis. We first argue that the use of the Pearson correlation for measuring the similarity between authors’ cocitation profiles is not very satisfactory. We then discuss what kind of similarity measures may be used as an alternative to the Pearson correlation. We consider three similarity measures in particular. One is the well-known cosine. The other two similarity measures have not been used before in the bibliometric literature. Finally, we show by means of an example that our findings have a high practical relevance.information science;Pearson correlation;cosine;similarity measure;author cocitation analysis
Application of CRISPR-based gene editing tools in Atlantic salmon
Get PDFAtlantisk laks er den viktigaste arten i norsk havbruksnæring. Næringa i Noreg er i dag ramma av fleire berekraftsufordringar, til dømes genetisk innblanding av rømt laks, og spreiing av smittsame sjukdommar. Dette påverkar både miljøet rundt, og fører til redusert velferd og helse hos fisken. Det er difor eit openbart behov for å beskytte oppdrettsfisken mot smittsame sjukdommar. Ei mogleg løysing på dette kan vere å endre fisken sin genotype ved bruk av genredigering, noko som kan bli gjort ved bruk av CRISPR/Cas genteknologi. Effektiv bruk av denne teknologien krev imidlertid evna til å uføre genredigering med høg nøyaktigheit. I tillegg trenger vi meir artsspesifikk kunnskap om kva gen som kan redigerast. Slik kunnskap kan vi få ved hjelp av CRISPR/Cas-baserte «loss-of-function» og «gain-of-function» eksperiment. Sagt på ein annan måte har CRISPR/Cas verdifulle bruksområde for både fiskeforsking og for havbruksnæringa.
Hovudmålet med denne avhandlinga var å implementere nye CRISPR-baserte verktøy i atlantisk laks. Dette målet er fokuset i Paper I og Paper II, kor vi testa CRISPR/LbCas12a genteknologi og base-redigering. Det sekundære målet var å bruke CRISPR-baserte verktøy til å lage in vivo modellar som kan bidra til å auke forståinga vår rundt biologiske prosessar knytt til laksen sitt immunsystem. Dette målet vart dekka i det siste manuskriptet, Paper III, kor CRISPR/Cas9 vart brukt til å slå ut dei to immungena Immunoglobulin (Ig) M og Interferon gamma (IFNg) i atlantisk laks.
Eksperimenta i denne avhandlinga har blitt utført ved å mikroinjisere fertiliserte lakseegg, etterfølgt av høgkapasitets-sekvensering for å undersøke resultata av genredigeringa. I Paper I vart CRISPR/LbCas12a brukt til å slå ut genet som kodar for pigmentering i laks, solute carrier family 45 member 2 (slc45a2), noko som resulterte i ein albino- eller albinomosaikk-pigmenteringsfenotype. Vi utførte òg eksperiment der donor templat med anten target- eller non-target DNA-tråd-orientering vart inkludert for å vurdere moglegheitene for å setje inn genetisk materiale med LbCas12a nukleasen. Her oppnådde vi opp til 54% integrerings-effektivitet. Til slutt injiserte vi både LbCas12a og SpCas9 CRISPR-kompleks i same individ for å samanlikne dei to nukleasane. Dei to produserte liknande mutasjonseffektivitet, men det var meir perfekt integrasjon av templatet på LbCas12a kuttstaden samanlikna med SpCas9 kuttstaden.
I Paper II brukte vi cytidin base-editoren AncBE4max for å introdusere eit stoppkodon i slc45a2. Vi oppnådde høg konverteringseffektivitet, opp til 89% C-til-T konvertering av den ønska mål-basen. I tillegg vart nokre uønska effektar oppdaga, som insersjonar og delesjonar, bystander-redigering, og konvertering av C-til-ikkje-T, men i liten skala. I Paper II vidareutvikla vi også eit alternativ til base-redigering, enkelt-base utbytting, ved bruk av konvensjonell CRISPR/Cas9. Her inkluderte vi templat med mutasjonar i ulike posisjonar frå kuttstaden. Her såg vi at integrerings-effektiviteten var påverka av mutasjonen sin avstand frå kuttstaden.
I Paper III vart CRISPR/Cas9 brukt til å slå ut IgM hos atlantisk laks, med meir enn 95% mutagenesis på begge IgM loci. Mutagenesis på IFNg mål-staden vart funne til å gjennomsnittleg vere ca. 57%, men med litt usikkerheit rundt dette. Utforskinga av effekten av å slå ut IFNg vart difor utsett i dette arbeidet, men er interessant for framtidige studiar. Effekten av å slå ut IgM på proteinnivå vart undersøkt ved hjelp av flow cytometry. I den genredigerte fisken var det gjennomsnittlege nivået av IgM positive (IgM+) B celler redusert med 91%.
For å konkludere har vi implementert to nye metodar for å utføre genetiske endringar i atlantisk laks, CRISPR/LbCas12 og base-redigering, og har dermed utvida verktøykassen for genredigering i denne arten. I tillegg brukte vi CRISPR/Cas9 til å lage ein F0 atlantisk laks med reduserte IgM+ B celle-nivå, som i framtida kan fungere som ein modell for å auke forståinga vår rundt dette genet, i tillegg til forståinga vår av den atlantiske laksen sitt immunforsvar generelt.Atlantic salmon is the most important species in the Norwegian aquaculture industry. The industry suffers from several sustainability challenges, such as genetic introgression of escapees and spreading of infectious diseases, which impact the surrounding environment and the welfare and health of the fish. Evidently, there is an overarching need to protect the farmed fish from infectious diseases. Changing the genotype of the fish by gene editing is a viable approach, and one powerful method to achieve this is by using CRISPR/Cas technology. However, effective application of this technology requires the ability to perform the edits with accuracy. Additionally, we need more species-specific knowledge of which genes to edit. Such knowledge can be obtained using CRISPR/Cas-based loss-of-function and gain-of-function approaches. Put differently, CRISPR/Cas has valuable applications for both basic fish research and the aquaculture industry.
The primary aim of this thesis was to implement new CRISPR-based gene editing tools in Atlantic salmon. This aim was the focus of Paper I and Paper II, where we tested CRISPR/LbCas12a technology and base editing. The secondary aim included using CRISPR-based tools to make in vivo models that can increase our understanding of biological processes related to the Atlantic salmon immune system. This aim is covered in the final manuscript, Paper III, where CRISPR/Cas9 was applied to knockout the immune genes Immunoglobulin (Ig) M and Interferon gamma (IFNg) in Atlantic salmon.
The experiments in this thesis have been performed by microinjecting fertilized salmon eggs, followed by high-throughput sequencing to reveal the editing outcomes. In Paper I, CRISPR/LbCas12a was used to knock out the pigmentation gene solute carrier family 45 member 2 (slc45a2) gene, resulting in an albino or albino mosaic pigmentation phenotype. We also conducted experiments where templates having a target or non-target strand orientation were added to evaluate the knock-in possibilities of LbCas12a, achieving up to 54% integration efficiency. Finally, LbCas12a and SpCas9 CRISPR complexes were injected in the same individual to compare the two nucleases. Interestingly, they both produced similar mutation rates, but more perfect integration occurred at the LbCas12a cleavage site compared to the SpCas9 cleavage site. In Paper II, the cytidine base editor AncBE4max was used to introduce a premature stop codon in slc45a2. We achieved highly efficient C-to-T conversion of up to 89% of our target base. Some undesired effects such as indels, bystander edits, and conversion of C-to-non-Ts were also observed but in small amounts. In Paper II we also further developed an alternative to base editing, single nucleotide replacement, using conventional CRISPR/Cas9. Here, templates featuring point mutations at various positions from the cleavage site were added, revealing that the insertion efficiency was affected by the mutation’s distance from the cleavage site.
In Paper III, we employed CRISPR/Cas9 to successfully knock out the two IgM heavy chain loci in Atlantic salmon, achieving more than 95% mutagenesis at both target sites. Mutagenesis on the IFNg target site(s) was found to be approximately 57%, although with some uncertainty. The investigations regarding IFNg were therefore postponed but may be interesting for future studies. The effect of the IgM knockout on protein level was assessed by flow cytometry analysis and revealed a mean average reduction in IgM positive (IgM+) B cells of 91% in peripheral blood of the gene-edited salmon.
In conclusion, we have implemented two new methods for making genetic changes in Atlantic salmon, CRISPR/LbCas12a and base editing, expanding the toolkit for gene editing in this species. We also employed CRISPR/Cas9 to generate a F0 IgM+ B cell-deficient Atlantic salmon, which may serve as a model to elucidate the role of this key gene and our understanding of the Atlantic salmon immune system in general.Doktorgradsavhandlin
Dispelling the Myths Behind First-author Citation Counts
Get PDFWe conducted a full-scale evaluative citation analysis study of scholars in the XML research field to explore just how different from each other author rankings resulting from different citation counting methods actually are, and to demonstrate the capability of emerging data and tools on the Web in supporting more realistic citation counting methods. Our results contest some common arguments for the continued
use of first-author citation counts in the evaluation of scholars, such as high correlations between author rankings by first-author citation counts and other citation
counting methods, and high costs of using more realistic citation counting methods that are not well-supported by the ISI databases. It is argued that increasingly available digital full text research papers make it possible for citation analysis studies to go beyond what the ISI databases have directly supported and to employ more
sophisticated methods
koamabayili/VECTRON-author-checklist: VECTRON author checklist
No full textWe have done our best to complete the author checklist relating to the use of animals in the hut study. Note that the objective for the hut study was to evaluate the IRS treatment applications for residual efficacy against Anopheles mosquitoes, including the local An. coluzzii mosquito population. Cows were only used to attract mosquitoes into the huts and no tests were carried out directly on the cows. The author checklist is intended for use with studies where experiments are carried out on animals, which is why we have had such difficulty in completing this for the hut study, as many of the questions do not relate to how the cows were used
Author-wise bibliometric analysis based on entropy.
No full textAuthor-wise bibliometric analysis based on entropy.</p
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