1,720,988 research outputs found
Amplicon clustering methods and the detection of core microbiota in honey bee gut
Get PDFRecent years decline in honey bee populations has lead to an increased interest in the study of their microbiota. In humans, a disturbance in the healthy gut microbiota is linked to several diseases, and because the host-adapted microbiota in the honey bee gut resembles that of mammals, it is assumed that bee gut microbiota also affects the health of bees. This leads us to the study of the core microbiota present in honey bee gut. Gaining knowledge about the core microbiota can help us understand what makes a healthy honey bee.
One route to acquire knowledge about the microbiota is by amplicon sequencing of the 16S rRNA gene. After sequencing, it is common to apply a clustering step to the data. Clustering methods can have a high impact on the results; the main focus in this thesis has therefore been to look at the effects of clustering methods in the study of amplicon reads.
Three clustering methods and a control method were used to group amplicon sequences to compare the differences of the clustering results and their ability to detect core microbes.
The results show considerable differences between methods, both in cluster composition and in the detection of core microbiota.
One of the clustering methods were not able to detect any core clusters (i.e., clusters part of every sample), and overlooked unique sequences present in a large number of samples.
Two methods did detect core microbiota, consistent with the core genera detected in previous studies. Besides, there were only detected minor differences in the core microbiota composition between different sampling factors such as time of year or gut part.
Results from this study clearly illustrate the importance of method when clustering amplicon reads. Depending on the choice of method, a study could end up with opposite conclusions regarding core microbiota
Tarmmikrobiotaen i den generelle populasjonen det første leveåret
Get PDFThe human gut microbiota consists of a complex community of microorganisms with
bacteria as the major resident. Their role in the gut can be viewed as an extension of
our genes and is often compared to a human organ. The gut bacterial community
follows us from birth and throughout our lives and has an implied role in maintaining
our health. Therefore, understanding factors that drive the early colonization is of
great importance. As most studies are based on selected cohorts, there is still a lack
of knowledge for the general population.
The aim of this thesis was to fill the knowledge gaps related to the establishment of
the gut bacteria, and the temporal changes during infancy in a general population. To
address this aim, we analyzed biological samples from a subset of the 2400 motherchild
pairs from the PreventADALL study. In total, we analyzed longitudinally
collected biological samples from 244 mother-child pairs. Fecal samples were
collected from mothers at 18 weeks of gestation, and their infants longitudinally (first
fecal sample, at 3 months, 6 months, and 12 months of age), in addition to skin swabs
of infants taken immediately after birth from 144 infants. These samples were
analyzed for different marker genes (16S rDNA and RpoB), metagenome sequencing
on subsets of infant samples (whole and reduced), in addition to metaproteomics and
metabolomics on infant sample subsets.
From our observations, the presence of Bacteroides in meconium, and on the infant
skin was dependent on birth mode. Vaginal born infants had Bacteroides as a
prominent member of the early gut community. Of the detected Bacteroides, two
maternal dominant strains were prevalent in the infant samples and persisted as a
gut bacterial member to at least 3 months of age. We identified a potentially globally
distributed B. caccae strain, and a potential local strain of B. vulgatus. Moreover, based
on the bacterial genomic content from the samples, diet seem to play a role in shaping
the early gut microbiota. The gut bacterial community’s potential ability for nutrient
utilization was skewed towards simple sugars, oligosaccharides, and mucins during early infancy, while this switched towards dietary components with the increasing
infant age.
The shift from breastmilk feeding to a diet consisting of solid foods was observed to
increase the abundance of Clostridium and decline the Bifidobacterium. The
combination of the diet- and bacterial shift was observed to have an effect on the fecal
short-chain fatty acid propionate, and butyrate. We observed two gut community
states during the transition from an infant- to an adult-like gut microbiota,
represented by high and low fecal butyrate. Within this cohort, we observed that E.
rectale and F. prausnitzii are potential key bacterial members for the increased
butyrate production, while infants with low fecal butyrate associated with R. gnavus.Tarmmikrobiotaen hos mennesker er sammensatt av et komplekst samfunn av mikroorganismer, hvor bakterier utgjør majoriteten. Rollen deres i tarmen kan bli sett på som en ekstensjon av våre gener, og blir ofte sammenliknet med et menneskelig organ. Sammensetningen av bakterier i tarmen følger oss fra fødselen og gjennom hele livet, og har blitt implisert i å spille en viktig rolle for å opprettholde vår helse. Derfor er det viktig å forstå faktorer som driver den tidlige koloniseringen. De fleste studier er basert på selekterte kohorter, og det er derfor mangel på kunnskap for den generelle populasjonen.
Målet for denne oppgaven var å undersøke etableringen av tarmbakterier, og følge denne utviklingen over tid i en generell populasjon. For å nå dette målet analyserte vi biologiske prøver på en delmengde av de 2400 mor-barn parene fra PreventADALL studien. Totalt analyserte vi biologiske prøver samlet kontinuerlig fra 244 mor-barn par. Avføringsprøver ble samlet fra mødre 18 uker inn i svangerskapet, og deres barn (den første avføringsprøven, ved 3 måneder, 6 måneder, og 12 måneder), i tillegg til svaberprøver tatt fra den nyfødtes hud rett etter fødsel av 144 spedbarn. Disse prøvene ble analysert for markørgener (16S rDNA og RpoB), metagenomsekvensering på en mindre del av barna (hel- og redusert), i tillegg til metaproteomikk og metabolomikk på noen utvalgte barn.acceptedVersio
Human gut microbiome : GA-map coverage and genomic categories in HumGut
Get PDFMicroorganisms in the gut are proven to affect health in multiple ways, and they have been studied extensively for the last decade. HumGut is a database containing microorganisms from the healthy human gut. The company Genetic Analysis (GA) has a GA-map dysbiosis test that aims to characterize microorganisms in the human gut to identify dysbiosis patients. This thesis provides an overview of the taxonomic and functional categorization of the human gut, gained by analyzing HumGut, and how well the GA-map spans the healthy human gut by investigating how it overlaps the database. Prodigal, Tax4FUN, and Diamond were used to acquire functional profiles for the HumGut genomes.
GA-map identifies microorganisms by matching GA-map probes to the 16S sequences in their genomes. Most genomes in HumGut do not have an identified 16S sequence. Still, the results show that the GA-map spans the majority of higher taxonomic ranks in HumGut. There are both taxonomic and functional parts missed by the map. The functional space missed by the GA-map seems to be relative similar to matched functional regions.
The genome categories in the HumGut collection are also evaluated in this thesis. The genomes in HumGut come from two sources: RefSeq and UHGG. Most of the UHGG genomes are Metagenome Assembled Genomes (MAGs). The findings show that MAGs may have lower quality on the 16S sequence than RefSeq-genomes, of which the latter is expected to have higher quality. The results also suggest that UHGG-genomes might have functional differences from other genomes.Det er bevist at mikroorganismer i tarmen påvirker helsen på flere måter, og det har blitt forsket mye på disse organismene det siste tiåret. HumGut er en database over mikroorganismer fra frisk human tarm. Bedriften Genetic Analysis (GA) har en GA-map dysbiose test med mål om å karakterisere mikroorganismer i human tarm for å identifisere dysbiose pasienter. Denne masteroppgaven presenterer en oversikt over human tarm gjennom taksonomisk og funksjonell kategorisering, oppnådd ved å analysere HumGut, og hvor godt GA-map spenner frisk human tarm ved å undersøke hvordan den identifiserer HumGutgenomene. Prodigal, Tax4FUN og Diamond ble brukt for å få funksjonelle profiler.
GA-map identifiserer mikroorganismer ved å matche med 16S-sekvensen til genomet. De fleste HumGut-genome har ikke en identifisert 16S-sekvens. Likevel viser resultatene at GAmap dekker over de fleste høyere taksonomiske nivåene i HumGut. Det er både taksonomiske og funksjonelle regioner som ikke dekkes. De funksjonelle regionene som ikke blir dekket av GA-kartet, ser ut til å være hovedsakelig like funksjonelle regioner som blir dekket.
En annen del av oppgaven er å evaluere genomkategoriene i HumGut. Genomene i HumGut kommer fra to kilder: RefSeq, som har validerte genomer, og UHGG. De fleste genomene i UHGG er Metagenom sammensatte genomer (MAGs). Funnene kan tyde på at disse genomene har lavere kvalitet på 16S sekvensene enn RefSeq-genomene, som er forventet å ha høyere kvalitet. Resultatene kan også indikere at UHGG-genomer kan ha funksjonelle forskjeller fra andre genomer.M-K
Bioinformatiske metoder for prediksjon av havbunnens økologiske tilstand fra Nanopore 16s rRNA gendata
Get PDFThe increasing anthropogenic pressures, particularly from industries and activities like intensive aquaculture affect marine benthic ecosystems. This context requires immediate implementation of frequent and effective environmental DNA monitoring methods. Technologies such as Nanopore sequencing provides an advanced and quicker analysis than the currently employed methodologies. However, there are various challenges, such as the need for an environment-specific database, that must be addressed to employ these monitoring strategies.
This thesis aims to mitigate these challenges and then predict the ecological state of seafloor samples collected across different ecological gradients. The study introduces METASEED, a novel method that combines both amplicon and metagenome data for reconstructing the 16S rRNA marker gene sequence, which has proven successful and has outperformed similar existing approaches (Paper 1). The METASEED and other existing approaches were implemented to generate a targeted database for environmental samples, consisting of 14,545 16S rRNA gene sequences (Paper 2). The comparative database analysis shows that strategies used for database creation are efficient in enhancing the AQUAeD-DB database diversity, producing consistent, positively correlated results for both Illumina and Nanopore read counts better than other database (median correlation coefficient: 0.50). The database also had more sea-floor associated taxa than other database since it was developed from seafloor samples.
The ecological status of seafloor sediments was predicted by implementing AQUAeD-DB database to assign Illumina and Nanopore reads using two different tools and three machine learning algorithms (Paper 3). The predictions based on Illumina and Nanopore data were comparable, showing similar prediction errors. Statistical analysis supported these findings, as there were no significant effects between the technologies used. There were significant effects between ML algorithms, with PLS outperformed Random Forest and LASSO. The feature selection using LASSO reduced the matrix dimension immensely and halved the prediction error. A high correlation was presented between observed and predicted nEQR values, with a Pearson correlation coefficient of 0.98 for Illumina and 0.95 (mean prediction error: ±0.04) for Nanopore data (mean prediction error: ±0.06).
This study concludes that in order to accelerate the benthic monitoring practices, Nanopore-based analysis can be a viable approach. However, it also underlines the necessity for standardizing the bioinformatic approaches and databases, which may need to be tailored for environmental samples to achieve optimal performance. Though here Nanopore provides comparable ecological prediction results to Illumina, it can still benefit from using Illumina as a reference.Det økende menneskeskapte presset, spesielt fra industrier og aktiviteter som intensiv akvakultur, pa virker de marine bentiske økosystemene. Dette krever implementering av raske og effektive eDNÅ-overva kingsmetoder. Teknologier som Nanopore-sekvensering tilbyr raskere analyse enn de na værende metodene. Det innebærer imidlertid nye utfordringer, for eksempel behovet for miljøspesifikke databaser.
Denne oppgaven adresserer disse utfordringene ved a bestemme den økologiske tilstanden til havbunnsprøver pa tvers av ulike økologiske gradienter. Oppgaven introduserer METÅSEED, en ny metode som kombinerer ba de amplikon- og metagenomdata for a rekonstruere 16S rRNÅ-gensekvenser (Årtikkel 1). METÅSEED og andre eksisterende metoder ble implementert for a generere en ma lrettet database (ÅQUÅeD-DB) for miljøprøver, besta ende av 14 545 16S rRNÅ-gensekvenser (Årtikkel 2). Vi utførte en sammenligning av ÅQUÅeD-DB og andre eksisterende databaser. Denne analysen viser at strategiene vi brukte til a lage ÅQUÅeD-DB forbedrer databasens artsmangfold, samt gir konsistente resultater for ba de Illumina og Nanopore signaler sammenlignet med andre databaser (median korrelasjonskoeffisient: 0,50).acceptedVersio
The GDR : a novel approach to detect large-scale genomic sequence patterns
Get PDFUtvikling av ny sekvenseringsteknologi de to siste tiårene har tillatt dypere dykk ned i de biomolekylære aspektene ved menneskets oppskrift. Hel-genom data fra flere hundre tusen mennesker er allerede tilgjengelig, men hvordan den økende mengden informasjon kan settes sammen til meningsfull funksjonell tolkning er komplisert og krever nye metoder. MikroRNA - mRNA interaksjoner utgjør et enormt genreguleringsnettverk som er vanskelig å predikere, selv for dagens beste maskinlæringsalgoritmer(1). Disse ikke-kodende elementene er involvert i omtrent alle cellulære prosesser i mennesket, primært via delvis komplementær baseparing mellom mikroRNA og mRNA, men det er mye vi ikke forstår av dette nettverkets betydning i vår biologi (2-4). Nye metoder er nødvendige for å kunne utforske genetisk variasjon i dette nettverket, som kan gi nye innblikk i hvordan genene våre reguleres.
Her presenteres «The Group Diversity Ratio» (GDR) som en ny målenhet til å møte denne utfordringen. GDR kan kvantifisere evolusjonær struktur av variasjon i store mengder genomisk sekvensdata, med et resultat som kan statistisk valideres. Metoden baserer seg på å måle gruppe-struktur i et distanse-basert fylogenetisk tre av sekvensdata, for forhåndsdefinerte grupper av «blader» i treet. Gruppene representerer en egenskap som kan relateres til sekvensdataen, og det undersøkes til hvilken grad det finnes en sammenheng mellom de to. Metoden kan primært brukes til å raskt skaffe overblikk over store mengder genomisk sekvensdata, som kan gi verdifulle innblikk til videre etterforskning.
For å teste metoden ble GDR brukt til å identifisere variasjon assosiert med etniske populasjoner i 3’UTR data fra «The 1000 Genomes Project» (1KGP). 1KGP var det første store prosjektet som adresserte den etniske skjevheten som nå finnes i genom-databaser, og som utgjør en god grunn til å utforske etnisk genetisk variasjon (5). I tillegg til identifikasjon av mer enn 1000 3’UTR sekvenser som inneholder signifikant etnisitet-spesifikk variasjon, viser dette studiet GDR-metodens høye potensial til å undersøke genetisk variasjon i stor skala
Benchmarking of metagenomic classification tools and storage stability of bioaerosol samples
Get PDFAir is a microbial habitat of crucial importance for public health. As such it is relevant for detection of potential epidemic or biothreat agents. The study of microbiological diversity in air through metagenomic analysis is a field under rapid development, and demands more knowledge. The work presented in this thesis investigated the current procedures used in metagenomic analysis of air samples, and consists of two parts. The first part assessed how long-time storage at low temperatures affects the stability of DNA concentration of filter-based air samples. Qubit and qPCR targeting the 16S rRNA gene were used to measure the DNA concentration. No evidence was found suggesting a detrimental effect of filter storage at – 80 °C. However, the findings may suggest negative effect of repeated freeze-thaw cycles on the yield of purified DNA. The second part assessed the performance of three metagenomic classification tools for creation of taxonomic profiles of air samples: Kraken 2, One Codex and Kaiju. The testing was conducted on various datasets. The results showed that Kraken 2 is the superior classification tool of well-studied species. However, Kraken 2 performed poorly on more complex datasets closer resembling the biological composition in air samples, due to inadequacies in the reference database. The classification of real air samples showed substantial variation between the profiles made by the tools. These findings further emphasise the need for improvements of the reference databases by adding more species specific for air, which should be a key objective for further work. There could also be improvements from altering the lowest common ancestor approach implemented in the classification algorithms, which seems to be a limiting factor for the taxonomic resolution
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
Genomanalyse av Salmonella enterica subsp. enterica serovar Typhimurium i Norge
No full textSalmonella enterica subspecies enterica serovar Typhimurium, referred to as S. Typhimurium in this master`s thesis, is one of the most common pathogenic serovariants of Salmonella spp. S. Typhimurium has been isolated in a wide range of hosts from the wild fauna. These hosts are believed to be responsible for several of the human cases of Salmonella infections in Norway. The main goal of this thesis is to test the hypothesis that wild animals are a potential source of infection from S. Typhimurium among production animals in Norway.
In this project a totalt of 127 S. Typhimurium isolates taken between 2001 and 2021 from wild animals, production animals and sport- and family animals were selected from the laboratory inventory at the Norwegian Veterinary Institute. The information present in the genome sequences of the S. Typhimurium isolates were examined using whole genome sequencing (WGS). WGS of pathogens is an important tool to improve the understanding of how contagious diseases spread between hosts. Due to limitations in the sequencing technology, the DNA are fragmented and only up to 300 bp are sequenced, this is then called reads. Bioinformatic tools were used for quality check a large number of the raw data from the sequencing before the original genom was attempted reconstructed using de novo SPAdes assembly.
” Multilocus Sequence Typing” (MLST) were used in further analysis to determine allele- variations based on 7 housekeeping genes, and each bacterial isolate were assigned one sequence type where ST19 and ST34 were the most common. Then the evolutionary similarity between isolates within the same ST was assessed by SNP analysis. Using phylogenetic analysis we were able to observe clusters with evolutionary similarity between wild animals and production animals, which indicates that transmission of S. Typhimurium had taken place. Based on the results from the applied dataset we found several of these cases, which gives a strong indication that wild animals can be the source of infection for S. Typhimurium among production animals in Norway
Comparative genomics of Salmonella enterica subsp. enterica serovar Typhimurium in Norway
Get PDFSalmonella enterica subspecies enterica serovar Typhimurium, referred to as S. Typhimurium in this master`s thesis, is one of the most common pathogenic serovariants of Salmonella spp. S. Typhimurium has been isolated in a wide range of hosts from the wild fauna. These hosts are believed to be responsible for several of the human cases of Salmonella infections in Norway. The main goal of this thesis is to test the hypothesis that wild animals are a potential source of infection from S. Typhimurium among production animals in Norway.
In this project a totalt of 127 S. Typhimurium isolates taken between 2001 and 2021 from wild animals, production animals and sport- and family animals were selected from the laboratory inventory at the Norwegian Veterinary Institute. The information present in the genome sequences of the S. Typhimurium isolates were examined using whole genome sequencing (WGS). WGS of pathogens is an important tool to improve the understanding of how contagious diseases spread between hosts. Due to limitations in the sequencing technology, the DNA are fragmented and only up to 300 bp are sequenced, this is then called reads. Bioinformatic tools were used for quality check a large number of the raw data from the sequencing before the original genom was attempted reconstructed using de novo SPAdes assembly.
” Multilocus Sequence Typing” (MLST) were used in further analysis to determine allele- variations based on 7 housekeeping genes, and each bacterial isolate were assigned one sequence type where ST19 and ST34 were the most common. Then the evolutionary similarity between isolates within the same ST was assessed by SNP analysis. Using phylogenetic analysis we were able to observe clusters with evolutionary similarity between wild animals and production animals, which indicates that transmission of S. Typhimurium had taken place. Based on the results from the applied dataset we found several of these cases, which gives a strong indication that wild animals can be the source of infection for S. Typhimurium among production animals in Norway.Salmonella enterica subspecies enterica serovar Typhimurium, omtalt som S. Typhimurium i denne masteroppgaven, er en av de vanligste patogene serovariantene av Salmonella spp. S. Typhimurium har blitt isolert hos et bredt spekter av verter fra villfaunaen som er antatt ansvarlig for flere av de humane smittetilfellene av Salmonella i Norge. Hovedmålet med oppgaven var å teste hypotesen om at ville dyr er en potensiell smittekilde for S. Typhimurium blant produksjonsdyr i Norge.
Det ble i dette prosjektet plukket ut totalt 127 S. Typhimurium isolater tatt mellom 2001 og 2021 fra ville dyr, produksjonsdyr og sport- og familiedyr fra laboratoriebeholdningen hos Veterinærinstituttet. Ved hjelp av helgenomsekvensering (WGS) ble informasjon som fantes i genomsekvensen til S. Typhimurium isolatene undersøkt. WGS av patogener er et viktig verktøy for å forbedre forståelsen av smittsomme sykdommer som sprer seg mellom verter. Begrensninger i sekvenseringsteknologien gjør at DNAet kuttes opp i kortere fragmenter, og kun korte fragmenter opp til 300 bp sekvenseres, også kalt reads. Ved hjelp av bioinformatiske verktøy ble store mengder rådata fra sekvenseringen kvalitetsjekket før det opprinnelige genomet ble forsøkt rekonstruert ved hjelp av de novo SPAdes assembly.
For videre analyser ble ” Multilocus Sequence Typing” (MLST) brukt for bestemmelse av allele-variasjoner basert på 7 husholdningsgener, og hvert bakterieisolat fikk tildelt en sekvenstype der ST19 og ST34 var mest vanlige. Deretter ble evolusjonær likhet mellom isolatene innenfor samme ST vurdert ved SNP analyse. For fylogenetiske analyser ble det observert klustre med en evolusjonær likhet mellom ville dyr og produksjonsdyr som indikerer smitteoverføring. Basert på resultatene fra det brukte datasettet var det i flere tilfeller sterke indikasjoner på at ville dyr kan være smittekilden for S. Typhimurium blant produksjonsdyr i Norge.M-K
Cost of resistance : stress tolerance and gene expression in wild type and mutant strains of Staphylococcus haemolyticus LMGT4071
Get PDFSeveral studies have shown that a bacteria’s development of resistance to an antimicrobial substance often comes with a cost related to their general fitness and adaptation to environmental changes. Further research is needed to increase the understanding on how bacteria with mutations in specific genes associated with stress tolerance, regulate their gene expression in response to stress.
The excessive use of antibiotics for treatment of pathogenic infections in patients have led to an increasing number of bacteria resistant to antibiotics. As it is crucial that we find alternative treatment methods, researchers are looking into the possibility of utilizing bacterial produced bacteriocins in combination with other antimicrobials. In the search for appropriate medicine, we need a broader knowledge of the bacteria’s response mechanisms related to the treatments.
Through this master’s thesis a mutant’s loss of fitness due to a resistance mutation will be examined. The study will attempt to give new inside on how a mutant of a bacterium regulates its gene expression when exposed to stress compared to the wild type within the same strain. Specific for this study, mutants within five bacterial strains with mutations connected to the function of the stress-response related gene rseP, are inspected in the laboratory. The bacteria were exposed to the bacteriocin Garvicin KS which do not use RseP as a receptor. RNA from samples of S. haemolyticus LMGT4071 will be isolated and sequenced to analyse the gene expression in wild type and mutant, with and without exposure to GarKS.
The experiments with S. haemolyticus in the laboratory show that the mutant tolerates the exposure to GarKS poorer than the WT, illustrating that the development of resistance comes with a fitness cost. The transcriptome analysis show that the mutant responds to the stress by altering its gene expression to a large extent, in contrast to the WT which turns on only a few genes.Flere studier har vist at en bakteries utvikling av resistens mot et antimikrobielt stoff ofte kommer med en kostnad knyttet til deres generelle fitness og tilpasning til miljøendringer. Det er behov for videre forskning for å øke forståelsen av hvordan bakterier med mutasjoner i spesifikke gener knyttet til stresstoleranse regulerer genuttrykket sitt i respons på stress.
Overdreven bruk av antibiotika for behandling av patogene infeksjoner hos pasienter har ført til at et økende antall bakterier er resistente mot antibiotika. Siden det er avgjørende at vi finner alternative behandlingsmetoder, ser forskerne på muligheten for å utnytte bakteriell produserte bacteriociner i kombinasjon med andre antimikrobielle midler. I jakten på passende medisin trenger vi bredere kunnskap om bakterienes responsmekanismer knyttet til behandlingene.
Gjennom denne masteroppgaven vil en mutants tap av fitness grunnet en resistensmutasjon undersøkes. Studien vil prøve å belyse hvordan en mutant av en bakterie regulerer genuttrykket sitt når det utsettes for stress sammenlignet med villtypen innenfor samme stamme. Spesifikt for denne studien inspiseres mutanter innenfor fem bakteriestammer med mutasjoner knyttet til funksjonen til det stress-responsrelaterte genet rseP i laboratoriet. Bakteriene ble eksponert for bacteriocinet Garvicin KS som ikke bruker RseP som reseptor. RNA fra prøver av S. haemolyticus LMGT4071 vil bli isolert og sekvensert for å analysere genuttrykket i villtype og mutant, med og uten eksponering av GarKS.
Forsøkene med S. haemolyticus i laboratoriet viser at mutanten tåler eksponeringen av GarKS dårligere enn villtypen, noe som illustrerer at utvikling av resistens kommer med en fitnesskostnad. Transkriptomanalysen viser at mutanten reagerer på stresset ved å endre genuttrykket sitt i stor grad, i motsetning til villtypen som kun slår på noen få gener.M-BIA
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