Identification of probiotic microbes from South African products using PCR-based DGGE analyses

[摘要] ENGLISH ABSTRACT:The regular consumption of probiotics is becoming a recognized trend in the foodindustry due to several reported health benefits. A probiotic is defined as a livemicrobial feed supplement that beneficially affects the host animal by improving itsintestinal microbial balance. A wide variety of probiotic food products are availableon the South African market and comprise an assortment of fermented milks, aswell as lyophilized preparations in tablet or capsule form.Strains of Lactobacillus acidophilus and Bifidobacterium species are mostlyused as probiotic microbes in the industry due to their health enhancing effect.The survival of sensitive probiotic microbial species in food matrices are influencedby various factors such as oxygen concentration, pH levels and manufacturing andstorage conditions. These should be considered and monitored as the SouthAfrican food and health regulations stipulate that probiotic microbes should bepresent 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 strainsand these factors have resulted in the need for the rapid and accurateidentification of probiotic microbes present in food products.The probiotic microbes present in probiotic yoghurts and supplements havein the past been identified using traditional methods such as growth on selectivemedia, morphological, physiological and biochemical characteristics. However,even some of the most sophisticated cultural-dependant techniques are not alwayssufficient for the identification and classification of especially Bifidobacterium, aswell as closely related Lactobacillus species. Molecular techniques are more oftenemployed for the rapid and accurate detection, identification and characterizationof microbial species present in food products.The aim of this study was to detect and identify the probiotic speciespresent in various commercial South African yoghurts and lyophilized preparationsusing peR-based DGGE analysis. A 200 bp fragment of the V2-V3 region of the16S rRNA gene was amplified and the peR fragments were resolved by DGGE.The unique fingerprints obtained for each product were compared to two referencemarkers A and B in order to identify the bands present. The results obtained wereverified by species-specific peR, as well as sequence analyses of bands thatcould not be identified when compared to the reference markers.Only 54.5% of the South African probiotic yoghurts that were tested didcontain all the microbial species as were mentioned on the labels of theseproducts, compared to merely one third (33.3%) of the lyophilized probiotic foodsupplements. Some Bifidobacterium species were incorrectly identified accordingto some product labels, while other products contained various microbes that werenot mentioned on the label. Sequence analysis confirmed the presence of apotential pathogenic Streptococcus species in one of the yoghurt products and insome instances the probiotic species claimed on the labels were non-scientific andmisleading.The data obtained in this study showed that the various South Africanprobiotic products tested were of poor quality and did not conform to the SouthAfrican regulations. peR-based DGGE analysis proofed to be a valuableapproach for the rapid and accurate detection and identification of the microbialspecies present in South African probiotic products. This could help with futureimplementation of quality control procedures in order to ensure a reliable and safeprobiotic product to the consumer.

[发布机构] Stellenbosch University