Thesis (MScFoodSc)--Stellenbosch University, 2004. / ENGLISH ABSTRACT:
The regular consumption of probiotics is becoming a recognized trend in the food
industry due to several reported health benefits. A probiotic is defined as a live
microbial feed supplement that beneficially affects the host animal by improving its
intestinal microbial balance. A wide variety of probiotic food products are available
on the South African market and comprise an assortment of fermented milks, as
well as lyophilized preparations in tablet or capsule form.
Strains of Lactobacillus acidophilus and Bifidobacterium species are mostly
used as probiotic microbes in the industry due to their health enhancing effect.
The survival of sensitive probiotic microbial species in food matrices are influenced
by various factors such as oxygen concentration, pH levels and manufacturing and
storage conditions. These should be considered and monitored as the South
African food and health regulations stipulate that probiotic microbes should be
present at a concentration of 10⁶ cfu.ml ̄ ¹' in order to exert a beneficial effect.
Some health benefits are also correlated to specific microbial species and strains
and these factors have resulted in the need for the rapid and accurate
identification of probiotic microbes present in food products.
The probiotic microbes present in probiotic yoghurts and supplements have
in the past been identified using traditional methods such as growth on selective
media, morphological, physiological and biochemical characteristics. However,
even some of the most sophisticated cultural-dependant techniques are not always
sufficient for the identification and classification of especially Bifidobacterium, as
well as closely related Lactobacillus species. Molecular techniques are more often
employed for the rapid and accurate detection, identification and characterization
of microbial species present in food products.
The aim of this study was to detect and identify the probiotic species
present in various commercial South African yoghurts and lyophilized preparations
using peR-based DGGE analysis. A 200 bp fragment of the V2-V3 region of the
16S rRNA gene was amplified and the peR fragments were resolved by DGGE.
The unique fingerprints obtained for each product were compared to two reference
markers A and B in order to identify the bands present. The results obtained were
verified by species-specific peR, as well as sequence analyses of bands that
could not be identified when compared to the reference markers.
Only 54.5% of the South African probiotic yoghurts that were tested did
contain all the microbial species as were mentioned on the labels of these
products, compared to merely one third (33.3%) of the lyophilized probiotic food
supplements. Some Bifidobacterium species were incorrectly identified according
to some product labels, while other products contained various microbes that were
not mentioned on the label. Sequence analysis confirmed the presence of a
potential pathogenic Streptococcus species in one of the yoghurt products and in
some instances the probiotic species claimed on the labels were non-scientific and
misleading.
The data obtained in this study showed that the various South African
probiotic products tested were of poor quality and did not conform to the South
African regulations. peR-based DGGE analysis proofed to be a valuable
approach for the rapid and accurate detection and identification of the microbial
species present in South African probiotic products. This could help with future
implementation of quality control procedures in order to ensure a reliable and safe
probiotic product to the consumer. / AFRIKAANSE OPSOMMING: Die gereelde inname van probiotiese produkte is besig om In erkende tendens in
die voedselindustrie te word, as gevolg van verskeie gesondheidsvoordele wat
daaraan gekoppel word. In Probiotika word gedefinieer as In voedingsaanvulling
wat uit lewendige mikrobes bestaan en wat In voordelige effek op mens of dier het
deur In optimale mikrobiese balans in die ingewande te handhaaf. In Wye
verskeidenheid probiotiese voedselprodukte is tans beskikbaar op die Suid-
Afrikaanse mark. Hierdie bestaan hoofsaaklik uit verskeie gefermenteerde
melkprodukte asook 'n reeks tablette en kapsules wat probiotiese mikrobes in
gevriesdroogde vorm bevat.
Lactobacillus acidophilus tipes en Bifidobacterium spesies word die
algemeenste in die voedselindustrie gebruik aangesien hierdie spesifieke
mikrobes bekend is om goeie gesondheid te bevorder. Die oorlewing van
sensitiewe probiotiese mikrobiese spesies in voedsel matrikse word beïnvloed
deur faktore soos suurstof konsentrasie, pH-vlakke en vervaardigings- en
opbergings kondisies. Hierdie faktore moet in aanmerking geneem word en
verkieslik gemonitor word aangesien die Suid-Afrikaanse voedsel en gesondheids
regulasies stipuleer dat probiotiese mikrobes teen In konsentrasie van 10⁶ kolonie
vormende eenhede per ml teenwoordig moet wees om In voordelige effek te toon.
Sommige gesondheidsvoordele word direk gekoppel aan spesifieke mikrobiese
spesies en spesie-tipes. Hierdie faktore het gelei tot In groot aanvraag na vinnige
en akkurate metodes vir die identifikasie van probioties mikrobes in
voedselprodukte.
Die probiotiese mikrobes teenwoordig in probiotiese joghurts en ook die
gevriesdroogde vorms in tablette en kapsules, was al geïdentifiseer deur gebruik
te maak van tradisionele metodes soos groei op selektiewe media, morfologiese,
fisiologiese en biochemiese eienskappe. Selfs van die mees gesofistikeerde
kultuur-afhanklike tegnieke is egter nie altyd voldoende vir die identifikasie en
klassifikasie van veral Bifidobacterium en na-verwante Lactobacillus spesies nie.
Molekulêre metodes word dikwels aangewend vir die vinnige en akkurate
deteksie, identifikasie en karakterisering van mikrobes teenwoordig in
voedselprodukte.
Die doel van hierdie studie was om die probiotiese mikrobes teenwoordig in
verskeie Suid-Afrikaanse joghurts en gevriesdroogde aanvullings, te identifiseer
deur gebruik te maak van polimerase kettingreaksie (PKR)-gebaseerde
denaturerende gradiënt jelelektroforese (DGGE) analise. 'n PKR fragment van
200 bp van die V2-V3 gedeelte van die 16S ribosomale RNS (rRNS) geen is
geamplifiseer, en die PKR fragmente is geskei met behulp van DGGE. Die unieke
vingerafdrukke wat verkry is vir elke produk is teen twee verwysings merkers A en
B vegelyk om die bande teenwoordig in die profiele te identifiseer. Die resultate is
bevestig deur spesies-spesifieke PKR en ook deur die ketting volgordes van die
DNS fragmente te bepaal wat nie geïdentifiseer kon word deur vergelyking met die
verwysings merkers nie.
Slegs 54.5% van die Suid-Afrikaanse probiotiese joghurts wat getoets is het
al die mikrobiese spesies bevat soos aangedui was op die etikette van hierdie
produkte, teenoor slegs 'n derde (33.3%) van die gevriesdroogde
voedingsaanvullings. Sekere Bifidobacterium spesies is verkeerd geïdentifiseer
op sommige van die produk etikette, terwyl ander produkte verskeie mikrobes
bevat het wat nie op die etiket aangedui was nie. 'n Potensiële patogeniese
Streptococcus spesie is in een van die joghurt produkte gevind soos bevestig deur
DNS kettingvolgorde bepalings. In sommige gevalle was die probiotiese
spesienaam wat aangedui is op die etiket onwetenskaplik en misleidend.
Die resultate wat uit hierdie studie verkry is dui aan dat die Suid-Afrikaanse
probiotiese produkte wat getoets is van 'n swak gehalte is en nie aan die Suid-
Afrikaanse regulasies voldoen nie. Daar is getoon dat PKR-gebaseerde DGGE
analise 'n waardevolle tegniek kan wees vir die akkurate deteksie en identifisering
van die mikrobiese spesies teenwoordig in probiotiese produkte. Dit kan help met
die toekomstige implementering van kwaliteitskontrolerings prosedures om 'n
mikrobiologiese betroubare en veilige produk aan die verbruiker te verseker.
| Identifer | oai:union.ndltd.org:netd.ac.za/oai:union.ndltd.org:sun/oai:scholar.sun.ac.za:10019.1/49983 |
| Date | 03 1900 |
| Creators | Theunissen, Johnita |
| Contributors | Witthuhn, R. C., Britz, T. J., Stellenbosch University. Faculty of AgriSciences. Dept. of Food Science. |
| Publisher | Stellenbosch : Stellenbosch University |
| Source Sets | South African National ETD Portal |
| Language | en_ZA |
| Detected Language | Unknown |
| Type | Thesis |
| Format | 71 p. : ill. |
| Rights | Stellenbosch University |
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