magistrsko delo
Leon Marič (Author), Janja Trček (Mentor)

Abstract

Iz industrijskih bioreaktorjev za proizvodnjo jabolčnega kisa smo v okviru projekta PKP osamili nove seve ocetnokislinskih bakterij. Analiza njihovih med genskih regij 16S-23S rDNA je odkrila tri seve (Komagataeibacter sp. AV382, Komagataeibacter sp. AV436 in Komagataeibacter sp. AV429), ki potencialno predstavljajo nove bakterijske vrste. V tem delu smo z različnimi fenotipskimi in genotipskimi pristopi pregledali njihove lastnosti z namenom, da ugotovimo njihov taksonomski položaj med ostalimi referenčnimi sevi ocetnokislinskih bakterij. V fenotipsko analizo smo uvrstili tudi vse referenčne seve iz rodu Komagataeibacter. Fenotipska preiskava je obsegala opazovanje morfologije kolonij, barvanje po Gramu, katalazni in oksidazni test, sposobnost sinteze celuloze in oksidacije ocetne kisline, identifikacijo posameznih vrst glukonskih kislin, analizo celokupnih maščobnih kislin, rast na različnih virih ogljika, rast v prisotnosti 30 % glukoze, sposobnost uporabe N iz (NH4)2SO4 ob prisotnosti različnih virov ogljika, rast brez etanola in ocetne kisline, rast ob prisotnosti 1 % ali 3 % etanola in različnih koncentraciji ocetne kisline ter rast v anaerobnih in aerobnih razmerah. Za sev Komagataeibacter sp. AV429 smo preverili tudi rast brez ocetne kisline, rast na acetatu pri pH 2,5, rast v prisotnosti etanola in laktata, rast na gojišču AE brez glukoze ter sposobnost uporabe različnih virov ogljika. Vsem sevom smo pregledali nukleotidna zaporedja genov za 16S rRNA in medgenske regije 16S-23S rDNA ter celotna genomska zaporedja. Ugotovili smo, da je genomsko zaporedje seva Komagataeibacter sp. AV382 najbolj podobno vrsti Komagataeibacter kakiaceti (ANIb 88,73 %) in seva Komagataeibacter sp. AV436 vrsti Komagataeibacter xylinus (ANIb 91,55 %). Na podlagi fenotipskih lastnosti lahko sev Komagataeibacter sp. AV382 razlikujemo od ostalih vrst rodu Komagataeibacter po tem, da producira 2-keto in 5-ketoglukonsko kislino, dobro raste na ribozi in propanolu, ne raste ob prisotnosti 30 % glukoze in ne sintetizira celuloze. Sposoben je uporabiti N iz (NH4)2SO4 v prisotnosti manitola na gojišču Hoyer-Frateur in Asai. V tekočem gojišču ne oblikuje biofilma, ob prisotnosti 1 % ali 3 % etanola pa tolerira do 5 % ocetne kisline. Sev Komagataeibacter sp. AV436 lahko na podlagi fenotipskih lastnosti od ostalih vrst rodu Komagataeibacter razlikujemo po tem, da sintetizira celulozo, tvori 2-keto in 5 keto-glukonsko kislino, dobro raste na ribozi in propanolu, vendar ne raste ob prisotnosti 30 % glukoze. Ob prisotnosti 1 % ali 3 % etanola tolerira do 5 % ocetno kislino. Sev Komagataeibacter sp. AV429 je po analizi genomske sekvence pokazal največjo podobnost z vrsto Gluconacetobacter entanii (ANIb 98,02 %), a njegove fenotipske lastnosti se od referenčnega seva znatno razlikujejo. Glede na zgornje rezultate predlagamo uvrstitev sevov Komagataeibacter sp. AV382 in Komagataeibacter sp. AV436 v novi vrsti znotraj rodu Komagataeibacter.

Keywords

magistrska dela;ocetnokislinske bakterije;kis;rod Komagataeibacter;taksonomija;

Data

Language: Slovenian
Year of publishing:
Typology: 2.09 - Master's Thesis
Organization: UM FNM - Faculty of Natural Sciences and Mathematics
Publisher: [L. Marič]
UDC: 663.318:663.15(043.2)
COBISS: 23994371 Link will open in a new window
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Downloads: 108
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Other data

Secondary language: English
Secondary title: Taxonomic Characterization of Acetic Acid Bacteria Komagataeibacter sp. AV382, Komagataeibacter sp. AV436 and Komagataeibacter sp. AV429
Secondary abstract: This study deals with novel strains of acetic acid bacteria isolated from industrial bioreactors for production of apple cider vinegar. The analysis based on 16S-23S rDNA resulted in identification of three strains (Komagataeibacter sp. AV382, Komagataeibacter sp. AV436 and Komagataeibacter sp. AV429) that could represent novel bacterial species. To determine their taxonomic position in comparison to other reference strains of acetic acid bacteria, we examined here their phenotypic and genotypic properties. The reference strains of Komagataeibacter genus have also been included in this study. The phenotypic analysis included colony morphology, Gram staining, catalase and oxidase activity, cellulose synthesis, acetic acid oxidation, detection and identification of gluconic acids, analysis of fatty acid methyl esters (FAME), growth on different carbon sources, growth in the presence of 30 % D-glucose, utilization of nitrogen from (NH4)2SO4, growth in the absence of ethanol and acetic acid, growth in the presence of 1 % or 3 % ethanol and different concentrations of acetic acid, and growth under anaerobic or aerobic conditions. For the strain Komagataeibacter sp. AV429 additional properties have been tested, such as growth in the absence of acetic acid on AE medium, growth on acetate at pH 2,5, growth in the presence of ethanol and lactate, growth on AE medium without glucose, and utilization of different carbon sources. For all three strains sequences of 16S rRNA gene, the internal transcribed spacer of 16S-23S rDNA, and whole genomic sequence have been analyzed. Results showed that the closest related species of Komagataeibacter sp. AV382 and Komagataeibacter sp. AV436 are K. kakiaceti (ANIb 88.73 %) and K. xylinus (ANIb 91.55 %), respectively. The strain Komagataeibacter sp. AV382 can be differentiated from other reference species of Komagataeibacter genus by its ability to form 2-keto and 5-keto-D-gluconic acid and incapability to grow in the presence of 30 % glucose. Furthermore, strain does not form pellet in liquid medium, and tolerates up to 5 % of acetic acid in the presence of 1 % or 3 % ethanol. It grows on Asai or Hoyer-Frateur medium supplemented with D-glucose or D-manitol and can utilize ribose and propanol as carbon sources. The strain Komagataeibacter sp. AV436 can be differentiated from other Komagataeibacter species by its abiliy to form 2-keto and 5-keto-D-gluconic acid, synthesize cellulose, and growth on ribose and propanol. Furthermore, in the presence of 1 % or 3 % ethanol, the strain tolerates acetic acid up to 5 %, but growth is absent in the presence of 30 % glucose. For the strain Komagataeibacter sp. AV429 genome sequence analysis showed a high similarity with strain Komagataeibacter entanii (ANIb 98.02 %), although their phenotypic properties differ substantially. Based on the data collected in this study, we propose for strains Komagataeibacter sp. AV382 and Komagataeibacter sp. AV436 classification into novel species of Komagataeibacter genus.
Secondary keywords: master theses;acetic acid bacteria;vinegar;Komagataeibacter genus;taxonomy;
Type (COBISS): Master's thesis/paper
Thesis comment: Univ. v Mariboru, Fak. za naravoslovje in matematiko, Oddelek za biologijo
Pages: IX, 82 f.
ID: 11601995