Use Of The RAD-PCR Fingerprinting For The Clustering Of Isolates Of Lactic Acid Bacteria From Fermented Milk (Rob) Samples

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Ahmed, Abdelhalim Abdullahi Hamza
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One hundred and sixty isolates of lactic acid bacteria were obtained from ten samples of Sudanese traditional fermented cow's milk (Rob), the most popular fermented milk in the Sudan. The samples were obtained from different locations in the Sudan. Thirty –two (18%) of the isolates were rod-shaped, and the rest (82%) were cocci. On the basis of a number of preliminary tests, which are commonly used for the identification of lactic acid bacteria, 42 homo- and heterofermentative representative isolates (14 rods and 28 cocci) were selected for further investigation and characterization. These were identified by utilizing morphological, physiological and molecular typing techniques. Randomly amplified polymorphic DNA (RAPD-PCR) fingerprinting using four oligonucleutide primers (A-11, C-4, O-10 and O-16) was the technique of choice for investigating the genetic diversity of the isolated strains. Fingerprinting the fourteen rod-shaped isolates resulted in five clusters based on numerical analysis of the RAPD-PCR profiles. Representative strains representing these clusters were identified by the API 50 STREP identification system. These were identified as Lactobacillus delbreuckii subsp .bulgaricus (one strain), Lactobacillus rhamnosus (9 strains), Lactobacillus plantarum (2 strains), Lactobacillus casei (one strain) and Lactobacillus pentosus (one strain). The selected 28 isolates representing the cocci were separated into nine clusters, the representative strains of which were identified by the API 20 STREP identification system. These representative strains were identified as Aerococcus viridans (4 strains), Enterococcus faecium (2 strains), Enterococcus gallinarum (2 strains), Lactococcus lactis subsp. lactis (5 strains), Leuconostoc sp (5 strains), Streptococcus acidominimus (8 strains) and Streptococcus bovis (2 strains). Some of the tested 42 strains produced exopolysaccharides from sucrose under the conditions used. These isolates can be utilized for the production of safe fermented milk products. Further studies on representative strains from these clusters can be put to use in the production of distinct rheologic and organoleptic traits in the production of Rob and other milk fermentation products. The present study demonstrates that the RAPD-PCR fingerprinting technique was more sensitive and reliable for differentiating between the isolates compared to the traditional and biochemical methods.
(Rob) Samples, Fermented Milk