Karakterizacija AggLb agregacionog faktora soja Lactobacillus paracasei subsp. paracasei BGNJ1-64
Characterization of AggLb aggregation factor from strain Lactobacillus paracasei subsp. paracasei BGNJ1-64
Author
Miljković, Marija S.Mentor
Kojić, Milan
Committee members
Jovčić, Branko
Lozo, Jelena

Kojić, Milorad
Kojić, Milan

Metadata
Show full item recordAbstract
Odgovor bakterija na uticaj razliĉitih faktora sredine koja ih okruţuje, kao i njihova
meĊusobna komunikacija, zavisi od procesa koji se odvijaju na površini bakterijskih ćelija. S
obzirom na brojne mogućnosti primene bakterija roda Lactobacillus, kako u prehrambenoj
industriji, tako i u medicini (u prevenciji i/ili leĉenju bolesti), potrebno je što bolje
okarakterisati komponente koje ĉine površinski sloj ovih bakterija. Agregacija bakterija,
determinisana faktorima lociranim na površini ćelija moţe se smatrati poţeljnom osobinom
pri izboru probiotika, s obzirom da ima vaţnu ulogu u meĊubakterijskoj interakciji, u
interakciji sa epitelijalnim ćelijama gastrointenstinalnog trakta i doprinosi sposobnosti
bakterija da se adsorbuju na razliĉite površine, odnosno sposobnost agregacije omogućava
uspešnu kolonizaciju staništa.
Kolekcija laktobacila Laboratorije za molekularnu mikrobiologiju je pretraţena na
prisustvo sojeva koji poseduju sposobnost agregacije (Agg+), nakon ĉega je za dalji r...ad
odabrano jedanaest sojeva: BGSJ2-8, BGGR2-68, BGGR2-82, BGDP1-84, BGDP9-38,
BGNJ1-3, BGNJ1-61, BGNJ1-64, BGNJ1-70, BGZLS30-6 i BGAR75. Odabrani Agg+ sojevi
se na osnovu stepena agregacije i tipa formiranih agregata mogu podeliti u tri grupe: oni koji
brzo, srednje i sporo agregiraju ili koji formiraju krupne, srednje i sitne agregate, pri ĉemu
postoji direktna korelacija izmeĊu brzine agregacije i tipa agregata. Analizom sposobnosti
vezivanja odabranih Agg+ sojeva laktobacila za komponente ekstracelularnog matriksa
(ECM) zapaţen je širok dijapazon sposobnosti vezivanja za kolagen - od intenzivnog afiniteta
vezivanja do odsustva sposobnosti vezivanja za matriks. Pritom su pojedini sojevi, BGGR2-
68 i BGGR2-82, ispoljili sposobnost vezivanja za plastiku (formiranje biofilma).
Analizom plazmidnog sastava Agg+ sojeva konstatovano je prisustvo velikih
plazmida u svim sojevima, a ĉišćenjem plazmida iz sojeva BGNJ1-64 i BGSJ2-8 potvrĊena je
plazmidna lokacija gena odgovornih za ekspresiju agregacionog fenotipa. Nakon konstrukcije
i analize plazmidnih biblioteka sojeva BGNJ1-64 i BGSJ2-8, izolovan je konstrukt pALb35
(koji nosi SacI fragment veliĉine 11,4 kb poreklom iz plazmida pNJ1 soja BGNJ1-64) koji
dovodi do ponovne pojave autoagregacionog fenotipa transformisanog heterologog domaćina
BGKP1-20 (Agg-)...
Response of bacteria to different environmental factors as well as their
communication primarily depends on the processes that occur on their cell surface. Due to the
continuously increasing number of application possibilities of lactobacilli, both in the food
industry and in medicine (in the prevention and/or treatment of the disease), there is a need
for a more comprehensive characterization of the components that form the surface layer of
these bacteria. Aggregation ability of bacteria, determined by factors located on the cell
surface could be considered as a desirable trait in the selection of probiotics. Since it plays an
important role in the interaction between bacteria, in interaction with the epithelial cells of
the gastrointestinal tract plus contributing to the bacterial capability to adhere to different
surfaces, which is an important feature enabling them to successfully colonize diverse
habitats.
Lactobacilli collection of Laboratory for molecular microbiology was screen...ed for the
presence of strains possessing the aggregation ability (Agg+) and for the further work eleven
strains have been selected: BGSJ2-8, BGGR2-68, BGGR2-82, BGDP1-84, BGDP9-38,
BGNJ1-3, BGNJ1-61, BGNJ1-64, BGNJ1-70, BGZLS30-6 and BGAR75 which, according
to the level of aggregation and the appearance of the aggregates formed, could be clasified
into three groups: the fast, the medium and the slow aggregating ones or into the groups of
those forming large, medium or small aggregates wherein a direct correlation between the
aggregation kinetics and the type of the formed aggregates was found. By analyzing the
binding ability of the selected Agg+ strains of lactobacilli to the components of extracellular
matrix (ECM) a wide range of binding capacity to collagen was observed, varying from the
intensive to the complete lack of the binding capacity to the matrix. Two strains (BGGR2-68
and BGGR2-82) showed the ability to bind even to plastics (form biofilms).
By analyzing the plasmid composition of Agg+ strains, the presence of large plasmids
was detected in all strains. The plasmid curing experiments confirmed the plasmid location of
the gene(s) responsible for the expression of aggregation phenotype in strains BGNJ1-64 and
BGSJ2-8. After the construction and analysis of BGNJ1-64 and BGSJ2-8 strains plasmid
libraries, selected construct pALb35 (which carries the SacI fragment of 11.4 kb. derived
from plasmid pNJ1 from strain BGNJ1-64) was able to reconstitute the autoagreggation
ability of the transformed heterologous host BGKP1-20 (Agg-)...