ITEM VIEW

Gastrointestinal persistence of the probiotic bacteria Lactobacillus plantarum 423 and Enterococcus mundtii ST4SA, and their anti-listerial activity

dc.contributor.advisorDeane, S. M.en_ZA
dc.contributor.advisorDicks, L. M. T.en_ZA
dc.contributor.authorVan Zyl, Winschau Fayghanen_ZA
dc.contributor.otherStellenbosch University. Faculty of Science. Dept. of Microbiology.en_ZA
dc.date.accessioned2018-07-23T09:19:23Z
dc.date.accessioned2018-12-07T06:46:25Z
dc.date.available2018-07-23T09:19:23Z
dc.date.available2018-12-07T06:46:25Z
dc.date.issued2018-12
dc.identifier.urihttp://hdl.handle.net/10019.1/104815
dc.descriptionThesis (PhD)--Stellenbosch University, 2018.en_ZA
dc.description.abstractENGLISH ABSTRACT: Gastrointestinal diseases, and in particular those caused by bacterial infections, are a major cause of morbidity and mortality worldwide. Treatment is becoming increasingly difficult due to the increase in number of species developing resistance to antibiotics. Different treatment strategies need to be developed. Probiotic lactic acid bacteria (LAB) have considerable potential as alternatives to antibiotics, both in prophylactic and therapeutic applications. Lactic acid bacteria have a long history of safe use in food and therapeutic products and is increasingly recognised for their beneficial effects. However, the underlying mechanisms by which probiotic LAB enhance the health of the consumer have not been fully elucidated. Demonstrating key antimicrobial and protective probiotic mechanisms in vivo will allow for industry and consumers to choose scientifically validated probiotics for the prevention or treatment of targeted gastrointestinal diseases. The present study aimed to contribute to this area of probiotic research. The first part of this study focus on monitoring the survival, metabolic activities and persistence of Lactobacillus plantarum 423 and Enterococcus mundtii ST4SA in the gastrointestinal tract (GIT) of mice using a bioluminescence imaging (BLI) system. The route and destination of both probiotic strains in the GIT were determined after single and multiple doses. Both strains prominently colonized the cecum and colon. Enterococcus mundtii ST4SA persisted in the GIT and faeces the longest and in higher numbers while also actively colonizing the small intestine. This is the first report of in vivo and ex vivo BLI of E. mundtii ST4SA in a murine model. The second part of the study encompassed the development of a novel system that facilitates the rapid and efficient isolation of double-crossover integration or deletion mutants of L. plantarum 423 and E. mundtii ST4SA. The system was useful in the construction of L. plantarum 423 and E. mundtii ST4SA bacteriocin and adhesion gene mutants. The newly described method expands the LAB molecular research genetic toolkit and has significant potential to allow genetic modification of most, if not all LAB species. This provides the unique opportunity to study the role of specific probiotic LAB genes in complex environments using reverse genetics analysis. In the final part of the study, the ability of L. plantarum 423 and E. mundtii ST4SA to competitively exclude L. monocytogenes EGDe, an intestinal pathogen, from the GIT of mice was proven. Valuable insight was gained on the molecular modes of action of the two probiotic strains. Plantaricin 423 and mundticin ST are bacteriocins produced by L. plantarum 423 and E. mundtii ST4SA, respectively. Bacteriocin-negative mutants of L. plantarum 423 and E. mundtii ST4SA failed to exclude L. monocytogenes EGDe from the GIT, confirming in situ bacteriocin production as an anti-infective mediator. Additional confirmation of in situ bacteriocin production as a mechanism of action was provided by using variant strains of L. monocytogenes EGDe expressing the immunity genes of plantaricin 423 and mundticin ST, respectively, which provided resistance to the respective bacteriocins. Furthermore, the exclusion of L. monocytogenes EGDe from the GIT was reduced when mice were administered with L. plantarum 423 and E. mundtii ST4SA adhesion gene knockout strains. These results substantiate our understanding of the functional attributes of probiotics currently available to consumers and the improvement of future probiotic products.en_ZA
dc.description.abstractAFRKAANSE OPSOMMING: Spysverteringskanaal (SVK) siektes, en veral dié wat veroorsaak word deur bakteriële infeksies, is wêreldwyd 'n belangrike oorsaak van morbiditeit en mortaliteit. Behandeling van SVK siektes word toenemend moeiliker weens 'n toename in die getal spesies wat weerstand ontwikkel teen antibiotika. Verskillende behandel strategieë moet ontwikkel word. Probiotiese melksuur bakterieë (MSB) het potensiaal as ‘n alternatief vir antibiotika, in profilaktiese- sowel as terapeutiese-toepassings. Melksuur bakterieë het 'n lang geskiedenis van veilige gebruik in voedsel- en terapeutiese produkte en geniet toenemend erkenning vir hul voordelige uitwerking op die mens se gesondheid. Die onderliggende meganismes waarvolgens probiotiese MSB die gesondheid van die verbruiker verbeter, is egter nie ten volle toegelig nie. Demonstrasie van belangrike antimikrobiese en beskermende probiotiese meganismes in vivo sal die industrie en verbruikers toelaat om wetenskaplik gevalideerde probiotika te kies vir die voorkoming of behandeling van geteikende SVK siektes. Die huidige studie beoog om by te dra tot hierdie gebied van probiotiese navorsing. Die eerste gedeelte van die studie fokus op die monitering van die oorlewing, metaboliese aktiwiteite en volharding van Lactobacillus plantarum 423 en Enterococcus mundtii ST4SA in die SVK van muise deur gebruik te maak van 'n bioluminisensie beeldvorming (BLB) stelsel. Die roete en ligging van beide probiotiese stamme in die SVK is suksesvol bepaal ná enkel- en veelvuldige dosisse. Beide stamme is prominent teenwoordig in die cecum en kolon. Enterococcus mundtii ST4SA het die langste, en teen hoër selgetalle, in die SVK oorleef, terwyl die dunderm ook aktief gekoloniseer is. Hierdie is die eerste bekendmaking van in vivo en ex vivo BLB van E. mundtii ST4SA in 'n muis model. Die tweede deel van die studie het betrekking op die ontwikkeling van 'n nuwe sisteem wat die vinnige en doeltreffende isolasie van dubbel oorkruis-integrasie of delesie mutante van L. plantarum 423 en E. mundtii ST4SA insluit. Die sisteem is nuttig vir die konstruksie van L. plantarum 423 en E. mundtii ST4SA bakteriosien- en adhesie mutante. Die nuut beskryfde metode maak ‘n belangrike bydrae tot die molekulêre tegnieke vir MSB en het die potensiaal om genetiese modifikasie van die meeste, indien nie alle, MSB-spesies, toe te laat. Dit bied die unieke geleentheid om die rol van spesifieke probiotiese MSB-gene in komplekse omgewings te bestudeer deur middel van omgekeerde genetiese analise. In die finale deel van die studie is bewys dat L. plantarum 423 en E. mundtii ST4SA die vermoë het om L. monocytogenes EGDe, ‘n patogeen, uit die SVK te verwyder. Waardevolle insig is verkry rakende die molekulêre werkswyse van die twee probiotiese stamme. Plantaricin 423 and mundticin ST is bakteriosiene wat onderskeidelik deur L. plantarum 423 en E. mundtii ST4SA geproduseer word. Bakteriosien-negatiewe mutante van L. plantarum 423 en E. mundtii ST4SA het nie daarin geslaag om L. monocytogenes EGDe in die SVK te onderdruk nie. Dit bevestig dat die in situ produksie van bakteriosiene ‘n belangrike rol speel in die bekamping van infeksies in die SVK. Ter bevestiging hiervan het stamme van L. monocytogenes EGDe, waarin die immuniteitsgene van plantaricin 423 en mundticin ST gekloneer is, weerstand gebied teen die twee bakteriosiene. Verdermeer het pogings om L. monocytogenes EGDe van die SVK te verwyder afgeneem met die dosering van L. plantarum 423 en E. mundtii ST4SA waarvan adhesie gene verwyder is. Hierdie resultate ondersteun ons begrip van die funksionele eienskappe van probiotika wat tans beskikbaar is vir verbruikers en die verbetering van toekomstige probiotiese produkte.af_ZA
dc.format.extent244 pages : illustrationsen_ZA
dc.language.isoen_ZAen_ZA
dc.publisherStellenbosch : Stellenbosch Universityen_ZA
dc.subjectLactic acid bacteriaen_ZA
dc.subjectProbioticsen_ZA
dc.subjectListeria monocytogenesen_ZA
dc.subjectUCTDen_ZA
dc.titleGastrointestinal persistence of the probiotic bacteria Lactobacillus plantarum 423 and Enterococcus mundtii ST4SA, and their anti-listerial activityen_ZA
dc.typeThesisen_ZA
dc.rights.holderStellenbosch Universityen_ZA


Files in this item

Thumbnail
Thumbnail

This item appears in the following Collection(s)

ITEM VIEW