Lactic Acid and Probiotic Bacteria

Lactic Acid and Probiotic Bacteria from Fermented and Probiotic Dairy Products
B.K.L. Karna1, O.C. Emata2 and V.L. Barraquio2*
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Institute of Agriculture and Animal Science, Central Campus, Rampur, Chitwan, Nepal Dairy Training and Research Institute, Animal and Dairy Sciences Cluster, College of Agriculture University of the Philippines Los Baños (UPLB), College, Laguna, 4031 Philippines Tel. No. (049)536-2203; Fax No. (049)536-2205 E-mail: vlbarraquio@yahoo.com Date submitted: July 18, 2006; Date accepted: January 11, 2007

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ABSTRACT
Lactic acid and probiotic bacteria were enumerated and isolated from commercially available yoghurt and probiotic milk products. Lactobacillus delbrueckii ssp. bulgaricus were enumerated and isolated using MRS agar incubated anaerobically at 37oC for 72 hrs. M17 agar was used for the enumeration and isolation of Streptococcus thermophilus incubated aerobically at 37oC for 48 hrs. MRS agar and modified MRS agar (MRS + L-cysteine + LiCl + Na propionate) were used for the enumeration and isolation of probiotic bacteria. Both were incubated anaerobically at 37oC for 72 hrs. Morphological, physiological and biochemical reactions were used to characterize the isolates. Str. thermophilus counts ranged from 2.6 x 1011 to 2.9 x 1020 CFU/g with Fruit Yoghurt (FY) having the highest count and Yoghurt Natural (YN) with the lowest count. Highest Lactobacillus delbrueckii ssp. bulgaricus count was obtained in Duo Yoghurt (DY), 1.1 x 109 and lowest in Yoghurt Drink (YD), 8.0 x 107 CFU/g. The highest probiotic bacterial count of 2.3 x 108 was obtained in Yakult (YK) and Neslac (Nes) showed the lowest, 1.6 x 102 CFU/g. The viable counts of all the products examined met the prescribed minimum viable count of 105 to 106CFU/g for the claimed health benefits for the consumer except for Chamyto Plain (CP), Nes and Nan-2 (Nan). Morphological, physiological and biochemical characteristics showed that the following genera and species were present Pediococcus acidilactici (YN), P. pentosaceus (FY), Lactobacillus delbrueckii delbrueckii and L. brevis in Non Fat High Calcium Yoghurt (NC), L. acidophilus and L. delbrueckii delbrueckii (DY, YD), P. damnosus and P. pentosaceus in Chamyto Orange (CO), L. delbrueckii bulgaricus, L. acidophilus, and L. delbrueckii delbrueckii (CP), L. para. paracasei (YK) and Bifidobacterium ssp. (Nes and Nan).Of the 28 isolates characterized in this study, 15 were Lactobacillus (5 species), 5 were Pediococcus (3 species), 6 were Bifidobacterium (species not identified), and 2 were Actinomyces israelii (1 species). Key words: LAB, lactic acid bacteria, probiotic, fermented dairy products

*Corresponding author

Science Diliman (July-December 2007) 19:2, 23-34

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Leuconostoc cremoris. L. L. 1988). L. adolescentis. which were all claimed to contain lactic acid bacteria (LAB). and absorption of calcium (David & Dauas. L. B. 1995). Lactobacillus acidophilus. The concept was introduced by Lilly & Stillwell (1965) and was intended to stimulate substances produced by one microorganism to enhance the growth of another. L. casei Shirota. plantarum. breve. casei immunitas. L. brevis. Their manufacturing date. MATERIALS AND METHODS Lactic Acid Bacteria (LAB) Locally available fermented dairy products. Metchnikoff first hypothesized the importance of lactobacilli for human health and longevity at the beginning of 19 th century. crispatus. johansonii. casei are widely used as probiotic bacteria in human and animal health. lechmannii. The health and nutritional benefits ascribed to probiotics can be generalized under the following categories: maintenance of normal intestinal microflora balance in infant and old age. B. Lactobacillus casei. The word probiotic was used later to refer to animal feed supplements and was defined as a live microbial feed supplement. antitumorogenic activity. L. acidophilus. Lactobacillus helveticus. L. especially during the last two to three decades to improve the health status by modulating the indigenous intestinal microflora by live microbial adjunct. 1989). 1998). Lactococcus cremoris. namely: Nestle Yoghurt Natural (YN). feeds. now called "probiotics" (Holzapfel et al. L. E. Lactococcus lactis. infantis. The products were collected from the supermarkets as fresh as possible. 1990). He considered the gut microbes as detrimental rather than beneficial and suggested that desirable effects might only be expected from their substitution by yogurt bacteria. and Duo Yoghurt (DY). The word "probiotic" was derived from the Greek word which means "on be half of ". cream and ripened butter and L. and Streptococcus sp. which are primarily used by the dairy industry. Probiotic therefore is the exact opposite of antibiotic. Non Fat High Calcium Yoghurt (NC). Anaerobic incubation was done using BBL Gas Pak (Code No. L. casei and L.. improvement of lactose tolerance and digestibility of the milk products.270308. Three (3) samples of each product were obtained and as much as possible. Number of colonies equal to the square root of the total colony count were isolated from the plates counted. 1998). Streptococcus thermophilus and Lactobacillus bulgaricus are used for manufacturing of yogurt. Fruit Yoghurt (FY). are used for the production of ripened cheese. The commonly used probiotic strains in different dairy products are: L. which beneficially affects the host animal by improving its intestinal microbial balance (Fuller. B. All samples were maintained at a temperature range of 4 to 60C prior to immediate microbiological examination. Lactobacillus sanfrancisco. Emata INTRODUCTION Lactic acid bacteria (LAB). L. animalis. B.Barraquio. reduction of serum cholesterol levels. Leuconostoc lactis. it was ensured that the triplicates were within the same or very close manufacturing dates. reuterii. Yoghurt Drink (YD). gasserei. cultured milk. Morphological characterization was done by 24 .. B. USA). delbrueckii. plantarum. Plating and isolation of LAB were done as shown in Figure 1 (IDF. B. 1989). lactis. 1991). Lactobacillus plantarum and Pediococcus acidilactici are used. The probiotic bacterial culture upon passage through the upper digestive tract must be capable of surviving and growing in the intestine and maintain their viability and activity in the carrier food before consumption (Gilliland. pharmaceuticals. Karna. and other applications. acidophilus and L. are extensively utilized in the fermentation of wide variety of food products and are known for their preservative and therapeutic effects (Gourama. faecium. B. This study deals with the isolation and identification of lactic acid and probiotic bacteria from locally available fermented and probiotic dairy products with the ultimate objective of preserving the isolates for their future potential use in the development of new fermented and probiotic foods. use by date and composition were carefully recorded. and E. Lactococcus diacetylactis. bifidum. brevis are used for the production of soda crackers (Gilliland. faecalis (Holzapfel et al. Becton Dickinson and Co. Since then attempts have been made. In meat and fish fermentation.. L. synthesis of B-complex vitamins. were used in this study as sources of LAB.longum.

examining colony growth. 69280 Marcy l'Etoile. thermophilus) anaaerobic (37oC/72 hrs) aerobic (37oC/48 hrs) Colonies Colonies isolation isolation MRS agar stabs Anaerobic (37oC/72 hrs) M 17 agar slants Aerobic (37oC/48 hrs) Figure 1. Biochemical characterization of the isolates was done following Holt et al. Non Fat High Calcium Yogurt (NC). 1988). 1998). Catalase test was done according to Harrigan & McCance (1976). The scheme for phenotypic identification is presented in Figure 2. 25 . France).Lactic Acid and Probiotic Bacteria LAB Fermented Dairy Products Yogurt Natural (YN). Gram reaction and cell size measurement (Benson. Fruit Yogurt (FY). Plating and isolation of LAB (IDF. bulgaricus) M 17 agar (Str. (1994) and using API 50 CH identification system (Biomeriux SA. Yogurt Drink (YD) and Duo Yogurt (DY) MRS agar (L.

Probiotic Bacteria Locally available probiotic dairy products from different supermarkets were used as the sources of probiotic bacteria. Fermented dairy products were the treatments to evaluate the presence of bacteria. Figure 2. For Chamyto and Yakult. Biotech. physiological and biochemical characterization were done similarly as for LAB.Barraquio.do - (D rd isca ) Bifidobacterium bifidus .01) and YN was the lowest.Switzerland) . 2. The probiotic dairy products and microorganisms claimed to be present in them are given in Table 1. Nes) Nan-2 (Infant dried milk. The reason for the 26 . 2. Yakult. Analysis of Variance (ANOVA) in CRD using the SAS soft ware was done on the data on bacterial counts. the diluents used were as recommended by IDF (1988).ve) G m -v ) ra ( e (Discard) (D rd isca ) Morphological. numbers and characteristics in the products (Gomez & Gomez. The diluents for Neslac and Nan were as specified in IDF (1999). Cabuyao. Manufacturer NestlePhilippines. UPLB. RESULTS AND DISCUSSION LAB and Probiotic Counts of Dairy Products The mean LAB and probiotic bacterial counts of different yogurt samples are given in Table 2.do - Gram G mea /n/Mrp o g reaction Morphology ra r ctio o h lo y L. All samples were maintained at a temperature range of 4 to 6o C during transit to the laboratory for immediate microbiological examination. Karna. The Least Significant Difference (LSD) test was used to determine differences among fermented dairy product means. Emata Stab/Slant Cultures Chamyto Plain (CP) Chamyto Orange (CO) S b/S n Cltues ta la t u r Dairy Products Probiotic Bacteria Claimed to be present Lactobacillus sp.do - Catalase (.9 x 1020 CFU/g (P ≤ 0. Phenotypic identification of LAB and probiotic bacteria Statistical Analysis Completely Randomized Design (CRD) with three replications per treatment was used. The scheme for plating and isolation of probiotic bacteria is presented in Figure 3. Japan) Table 1 Probiotic dairy product samples Gram (+ ve) G m +v ) ra ( e Gram (.. Colonies were picked from the plate such that the number of isolates taken was equal to the square root of the total colony count.ve) Cta se(-v a la e) Catalase (+ ve) Cta se(+v a la e) (Discard) Neslac (Infant dried milk. their types. API test (Sugar Fermentation) AIte (S g rF e ta n P st u a ermn tio ) Figure 2. use by date and composition were carefully recorded. thermophilus counts of FY was highest. Nan Yakult Fermented milk. Laguna. YK) Bifidobacterium bifidus .Their manufacturing date.do - . (Nestle.6 x 1011 CFU/g. casei strain Shirota Yakult Philippine Inc. 1984). Identified LAB and probiotic bacteria were preserved as described by Kisworo & Barraquio (2003) and deposited at the Philippine Network of Microbial Culture Collections (PNMCC). Laguna (Collab. The data showed that the mean Str.

30 x 108 CFU/g and Nes showed the lowest. 1999) higher mean putative count of FY may be due to the differences in manufacturing dates of samples used. Liquid milk has an aw of about 0. NC. Plating and isolation of probiotic bacteria (IDF. while YD was lowest. The mean probiotic count of Nan (1 year-old) was significantly higher than the Nes (2 27 . The mean Lactobacillus delbrueckii ssp. FY and YN were statistically at par with each other but significantly higher than the YD. 8.00. aw. 1.Highest Lactobacillus delbrueckii ssp. The data showed that the mean counts of DY. which may be due to the loss of nutrients in the yogurt and accumulation of waste with age (three weeks) of the sample (Kozaki et al.) (for Bifidobacterium) MRS agar (for LAB) Anaerobic 37 0 C / 72 hrs Table 1 Probiotic dairy product samples Anaerobic 37 0 C / 72 hrs Colonies Colonies Modified MRS stabs MRS stabs Anaerobic. The reason for the differences in mean counts may be due to the nature of the product. Probiotic bacterial count of YK was 2. 37 0 C / 72 hrs Figure 3.98 while dried or powdered milk's aw is below 0. This water requirement is best expressed in terms of available water or water activity. The amount of water needed for growth of microorganisms varies. bulgaricus counts of different yogurt samples are also given in Table 2. 1992).05). bulgaricus count was obtained in DY. Nes being a powder. Most bacteria grow well in a medium with aw approaching 1. 1988).6 x 102 CFU/ g. The data also showed that the mean putative probiotic count of YK was significantly higher than the other samples.0 x 107 CFU/g (P ≤ 0..Lactic Acid and Probiotic Bacteria Probiotic Dairy Products Neslac and Nan-2 Chamyto and Yakult Modified MRS agar (MRS +L-cysteine +LiCl +Na-prop. 1.1 x 109 CFU/g.60 (Frazier and Westhoff.

6 x 103 d 1. The LAB belonging to the genus 2. DY-2. Nes having earlier manufacturing date. delbrueckii ssp. Nan-1.1967). The morphological characteristics of LAB and probiotic isolates are presented in the Table 4. Pediococcus as described by Sneath et al (1986). Nes and Nan did not meet the prescribed minimum viable counts mentioned. thermophilus and 72 hr-old culture in case of L. bulgaricus and probiotic bacterial isolates. for the consumers to obtain the claimed health benefit. transferred to stab/slant for study of the growth pattern of isolates on the solid media.1 x 109 a 2. YK-7.bulgaricus). CO-5 and CO-6 have colony characteristics that resembled the genus.6 x 102 e Means with the same superscript are not significantly different at P ≤ 0. well isolated and representative colony that appeared on the plate. irregular and arborescent on slant streak and stab.4 to 1. Shah (2000) recommended a minimum LAB count of not less than 106 CFU/g in fermented dairy product. triangular. The colonies appeared grayish white in color. DY-6.0 to 4. five isolates namely.. YD-1.9 x 1020 a 1.6 x 1014 c 1. Nan-5 and Nan-6.0 by 2 to 10 µm whereas the cell measurements of Pediococcus isolates ranged from 1. CP. six (6) isolates namely. Isolates Nes-2 and Nan3 resembled Actinomyces.1 x 108 a 1. Plain (CP) Nan . Regarding probiotic bacteria. filiform. smooth and regular growth on stab. YD-2. soft to mucoid which also resembled more or less Bifidobacterium (Sneath et al. Colony Characteristics of LAB and Probiotic Bacterial Isolates Colony characteristics of LAB and probiotic isolates were studied by picking up the typical.thermophilus) and P ≤ 0. thermophilus1 2. NC-6.4. Nan-4. showed colony characteristics that resembled Bifidobacterium sp. smooth entire and filiform. Cell measurements of isolates were done under the high power objective (400 x). bulgaricus1 2.0 µm in diameter. smooth. round. Emata years old). (1994) described the colonies as creamy white glistening with soft consistency. round. Nes-1.05 (L. Cell measurements were based on 48 hr-old culture in case of Str.01 n = number of samples examined Table 2 Mean LAB and probiotic bacterial counts 28 . triangular and spindle shaped. round. CP-5 and CP-6 were found to be typical of genus Lactobacillus colonies of which generally appeared white to yellowish in color. the minimum number of LAB and probiotic bacteria should be maintained at 105 CFU/g of cultured product. YK-2.6 x 108 a 8. YN-6. A single colony was aseptically picked-up and SAMPLE For LAB MEAN COUNTS (n=3. In this study. YK-3. CP-4. round. star-like structure with effuse. 1986. CFU/g) Str. raised and convex elevation.3 x 108 a Morphological Characteristics of LAB and Probiotic Bacterial Isolates The LAB and probiotic isolates were characterized morphologically by Gram staining. Holt et al. DY-3. spindle. Waksman. FY.0 x 107 b Fruit Yogurt (FY) Yogurt Drink (YD) Non fat high Ca Yogurt (NC) Duo Yogurt (DY) Yogurt Natural (YN) For Probiotic bacteria2 Yakult (YK) Chamyto. which were creamy white in color. which may be due to the age of the sample.8 x 1013 d 2. All 70 isolates were Gram positive. Nan-2. According to Robinson (1987). 2 Means with the same superscript are not significantly different at P ≤ 0. Karna.6 x 106 b 3. FY-5.4 x 104 c 1. Orange (CO) Chamyto. DY-1.01 (S.6 x 1011 e 3.3 x 108 a 3. YK-1. Fifteen isolates namely.2 (Nan) Neslac (Nes) 1 2.4 x 1019 b Lacto.Barraquio.4. Cell measurements of LAB isolates ranged from 0. Growth of isolates was observed and cultural characteristics were described (Table 3). NC.

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 ISOLATES YN . NC-6. acidophilus Lact. very small size. raised regular & punctiform growth on plates & filiform and irregular growth on stab Colonies triangular. Actinomyces israelii Bifidobacterium sp.delb Lact. regular & entire and slant growth is filiform Colonies round. punctiform & beaded type growth on stab Colonies round.2 x 1.acidilactici Lact. brevis Pedio. raised. surrounded by transparent area on solid media and punctiform on slant.delb.5 to 1. Lact acidophilus Lact.paracasei Lact. white on solid media. Pedio. non transparent. convex. arborescent and irregular on stab Colonies triangular. DY-1.delb. Bifidobacterium sp. namely YN-6.. (1994). YK-3. arborescent and irregular growth on stab Colonies triangular. CP-4. entire and filiform growth on stab. Bifidobacterium sp. cream color. YK-7. smooth.delb delb Lact. Five (5) isolates.. DY-6. light yellow. convex. raised. usually regular but sometimes they are almost coccoid. delb. smooth & regular growth on stab Colonies small dot structure.delb. damnosus Pedio pentosaceus Lact. small. Colonies round. smooth. spindle. COLONY CHARACTERISTICS Colonies round.star like on plate and arborescent and irregular on stab Colonies small. ( ) not a LAB nor probiotic bacteria Table 3. YD-2. raised and entire growth on slant Colonies round. commonly in short chains with cell size of 0. white. entire and arborescent growth on stab.Lactic Acid and Probiotic Bacteria Lactobacillus have rod shaped cells. convex.6 FY-4 F L -5 DY -1 DY-2 DY. Colony characteristics of LAB and probiotic bacteria Table 3. cream color. white raised and non-transparent filiform. 1994). convex & non transparent. regular and dot like on plate. smooth and filiform growth on stab Colonies round. CP-5 and CP-6 resembled the genus Lactobacillus as described by Holt et al.0 µm in diameter in pairs or tetrads (Holt et al. entire and filiform growth on stab.4 NC . regular. convex. Out of 28 isolates.delb bulg. YD-1. DY-2. raised. regular. raised and effuse growth on solid media. light yellow. FL-5.pentosaceus Pedio pentosaceus Lact. Colony characteristics of LAB and probiotic bacteria 29 . smooth white and flat on plate the growth is arborescent. raised and entire growth on slant Colonies round. regular. acidophillus Lact. raised. white glistening convex soft on plate and arborescent growth on stab Colonies spindle.0 to 2. FL-4. Colonies round. raised. convex. cream color. smooth and cream color & filiform growth on stab Colonies round creamy. flat and yellow color and effuse growth on solid media. regular. regular.white raised on plate and arborescent and irregular growth on stab Colonies round. Bifidobacterium sp. Colonies round.paracasei Lact. flat. Actinomyces israelii Bifidobacterium sp. convex. raised and filiform and entire growth on solid media Colonies round. yellow flat and filiform. YK-1. Colonies spindle. Colonies spindle. were found to be typical of the genus NO. white glistening. transparent. paracasei Lact. white. paracasei Lact. fifteen (15) isolates namely: NC-4. 1. acidophillus Lact. white. white and rose Colonies round.6. CO-5 and CO. non-transparent.delb. convex. and punctiform on plate & filiform growth on stab Colonies yellow. smooth. entire.. raised.3 DY -6 YD-1 YD-2 CO-5 CO-6 YK-1 YK-2 YK-3 YK-7 CP-4 CP-5 CP-6 Nes-1 Nes-2 Nan-1 Nan-2 Nan-3 Nan-4 Nan-5 Nan-6 IDENTITY Pedio. white raised& smaller size effuse and entire growth on slant Colonies round. DY-3. Bifidobacterium sp.0 to 10 µm while the genus Pediococcus has spherical cells.irregular.6 NC . YK-2. entire and arborescent growth on stab Colonies triangular. entire and white filliform. entire. raised and filiform entire growth on solid media Colonies round. smooth. white. transparent. flat on plate arborescent and irregular growth on stab Colonies round. surface growth on broth and arborescent growth on stab Colonies round. acidophilus Lact. smooth & entire and arborescent growth on stab Colonies spindle.

x 2.6-1. brevis Lact.bulgaricus Lact. Another work conducted by Tzanetakis and Litopoulou-Tzanetaki (1992). lactis. Rodrigues et al. some are in pairs Cells rod shaped. in tetrads. delb.0 µm 1. in tetrads. in case of LAB.0 x 2-4 µm 0. with square ends and in chains Cells thin rods. Nes-1. tetrads. Sugar Fermentation of LAB and Probiotic Bacterial Isolates Out of 70 isolates. casei followed by Lactococcus lactis ssp. The summary of API output of LAB and probiotic bacterial isolates is presented in Tables 6 and 7. Waksman (1967) explained that Actinomyces occurs virtually in every natural substrate such as fresh water basins. The sugar fermentation patterns of LAB and probiotic bacterial isolates are presented in Table IDENTITY Pediococcus acidilactici Pediococcus pentosaceus Pediococcus damnosus Lact.4.5-1.5-8 µm 0. branched. which comprised fourteen (14) out of twenty-three (23) isolates.5-1.0 µm 1. Nan-4. Karna. only 28 isolates were selected and subjected to sugar fermentation test due to budgetary constraints. (1995).1.3 x 1. acidophilus Lact.4-1. showed that Lactobacillus was the predominant genus over Enterococcus and Pediococcus in cheese. delb. branched. delbrueckii Lact. cell shape. para. majority of the isolates were Lactobacillus with 5 different species identified followed by Pediococcus with 3 different species. while the sources of LAB and probiotic isolates are shown in Table 8. Actinomyces israelii was also isolated from Nes-2 and Nan-3 which might have come from the processing plant environment. Cells rod shaped. in his study on the biodiversity of LAB from food related ecosystem also reported that the Lactobacillus strain dominated all ecosystems and consisted 65% of Lactobacillus isolates.0 . foodstuffs and the atmosphere and multiply most abundantly in various depths of soil and compost in temperate and tropical regions. (Actinomyces israelii) SHAPE Cells spherical. (1994). Nan-2. Nan-5 and Nan-6 appeared as thin. regular. cell size and occurrence.0 µm 0.2. v and y arrangements in chains.6 µm 0.5-1. occur in pairs or chains with square ends Cells thin rods.0 x 2-8 µm 0.2. Damelin et al.Barraquio. namely. also in pairs Cells spherical. arranged singly and pairs in V and Y arrangements.0 . Gram positive rods.4µm ( ) not a LAB nor probiotic bacteria Table 4 Morphological characteristics of LAB and probiotic bacterial isolates 30 . Six (6) of the probiotic isolates.0 x 4-8 µm 0.0 . reported that majority of species found in raw cow milk cheese were Lactobacillus casei ssp. Nan-1. occurring in short chains Cells rod shaped. 5.4 . Kisworo and Barraquio (2003). Cells short rods with clubbed ends occurring in v and y arrangements in chains AVERAGE CELL SIZE 1.0 x 4 . in chains.0 x 4 -10 µm 0. regular. In general. typical of Bifidobacterium as described by Holt et al. (1995). also pairs Cells spherical. also found in their study of raw milk and white soft cheese that the most predominant genus isolated was Lactobacillus. occur singly/chains Cells rod shaped. regular rounded ends. Emata Pediococcus in Gram reaction. respectively.6-1. occurring in short chains. paracasei Bifidobacterium sp.

Lactobacillus brevis 7 -Pediococcus damnosus 9 -Bifidobacterium spp. delbrueckii 6.Lactic Acid and Probiotic Bacteria ISOLATES SUGAR Glycerol L-arabinose D-arabinose Ribose D-xylose D-glucose D-fructose D-mannose L-sorbose Sorbitol Rhamnose N-A.arbitol Gluconate Adonitol Indol Urease 1 + + + + + + + + + + + + + 2 + + + + + + + + + + + + + + + + + + + + + + + + 3 + + + 4 + + + + + + + 5 + + + + + + + + + + + + + + + + + + + + + + + + 6 + + + + + + + + + + + + + + + + 7 + + + + + + + + + + + + + + 8 + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + 9 + + 10 + + 1.Lactobacillus acidophilus 3 -Lactobacillus delb.D-glucoside Galactose Gelatin Trehalose Cellobiose Saccharose Melezitose Inulin D-raffinose B-gentiobiose D-turanose D-tagatose D.Lactobacillus para. Sugar fermentation patterns of LAB and probiotic bacterial isolates 31 . bulgaricus 5. 2.Actinomyces israelii Table 5.paracasei 4 -Lactobacillus delb.glucosamine Amygdalin Arbutine Esculin Salicine Manitol Maltose Lactose α-M.Pediococcus pentosaceus 8 -Pediococcus acidilactici 10.

9 99. brevis Pedio. acidophilus Lact. delb. 01 02 03 04 05 06 07 08 09 10 11 12 13 14 15 16 17 18 19 20 Isolates Y N-6 NC -4 NC-6 FY -4 FY -5 DY -1 DY . Lactobacillus para. Bifidobacterium sp. bulgaricus Lact. Bifidobacterium sp. acidophilus Lact.5 CP .3 YK .7 85.7 99. paracasei Lact. para.4 CP .9 99. damnosus Pedio.delbrueckii Pedio. Lactobacillus acidophilus and Lactobacillus delbrueckii delbrueckii (CP).3 DY .9 99. paracasei (YK). pentosaceus Lact. (Actinomyces israelii) Bifidobacterium sp. and Bifidobacterium sp. physiological and biochemical characteristics showed that the following genera and species of LAB and probiotic bacteria were present in the dairy products examined: Pediococcus acidilactici (YN).Barraquio.5 % 95.delbrueckii Lact. Lactobacillus delbrueckii bulgaricus. delb.delbrueckii LAB and Probiotic Bacteria Pedio.1 99.delb. delbrueckii Lact. acidophilus Lact. Pediococcus pentosaceus (FY). five (5) were Lactobacillus acidophilus. Emata No.2 CO-5 CO-6 YK .delbrueckii Lact.acidophilus Lact. pentosaceus Lact.6 YD . delb. delb.9 98. (Actinomyces israelii) Total isolates ( ) not a LAB nor probiotic bacteria Yakult (plain) Neslac (powder milk) Nan-2 (powder milk) Table 6 Summary of API identification output for LAB Table 8 Sources of LAB and probiotic bacterial isolates No.9 96. para.8 99. acidophilus Lact. four (4) each of L. paracasei Lact.7 98.5 99. paracasei Lact. acidophilus Lact. (Actinomyces israelii) Bifidobacterium sp. pentosaceus Lact.2 YK . delb. Lactobacillus delbrueckii delbrueckii and Lactobacillus acidophilus (YD).1 YD . para.delb.2 % 94. morphological. Bifidobacterium sp. acidophilus Lact.6 IDENTITY % Identification 99. para. Lactobacillus delbrueckii delbrueckii and Lactobacillus brevis (NC).2 % 96. Karna. Of the twenty-eight (28) isolates.7 CP . acidophilus Lact.8 99. Pediococcus damnosus and Pediococcus pentosaceus (CO). of Isolates 1 1 1 2 1 3 1 1 1 1 1 1 1 4 1 1 5 1 28 Non Fat High Ca yogurt Lact.2 95. pentosaceus Lact.7 % 96.delbrueckii Pedio.1 99.pentosaceus Pedio. paracasei Lact.8 99. ( ) not a LAB nor probiotic bacteria Table 7 Summary of API identification output for probiotic bacterial isolates 32 . Lactobacillus acidophilus and Lactobacillus delbrueckii delbrueckii (DY). damnosus Pedio. delbrueckii Lact. paracasei Bifidobacterium sp.5 % SUMMARY AND CONCLUSIONS Cultural.7 99. brevis Fruit yogurt Duo Yogurt Yogurt drink Chamyto (orange) Chamyto (plain) Pedio.2 % 90. 1 2 3 4 5 6 7 8 Isolates Nes-1 Nes-2 Nan-1 Nan-2 Nan-3 Nan-4 Nan-5 Nan-6 IDENTITY Bifidobacterium sp. % Identification 98.2 DY .9 90.9 99. delb delb rueckii Lact.2 % 95.5 92. bulgaricus Lact.delb.para. acidilactici No.delb. (Actinomyces israelii) Bifidobacterium sp.2 Sources Yogurt Natural Pediococcus acidilactici Lact. (Nes and Nan).1 YK .5 % 95.

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