According to propionibacteria natural habitats, nine species have been recognized to belong to the genus Propionibacterium and grouped to dairy propionibacteria. These strains were comprising Propionibacterium thoenii (3), Propionibacterium acidipropionici (2), Propionibacterium freudenreichii ssp. shermanii (1), Propionibacteriun freudenreichii (1), Propionibacterium freudenreichii ssp. freudenreichii (1) and Propionibacterium shermanii (1). All nine strains were subjected to in vitro analysis to assess their probiotic potential. Eight strains of Propionbacterium were fully tolerant to 0.3% and 0.5% bile salts and were able to survive at pH 3 for 3hrs while at pH 2 a few strains were able to survive. Moreover, good growth was observed in the presence of 0.3% phenol. Antimicrobial activity of Propionibacterium strains was found to be good against most of tested strains. In addition, all tested strains were sensitive to tertracyclin and trimethoprime/ sulfamethoxazole. Technological and productive characteristics tests showed that tested strains disaplayed various behaviours in their acidifying activity and four strains produced exopolysaccharide. In conclusion, eight strains P. thoenii P15, P. thoenii TL18, P. freudenreichii ssp. shermanii ATCC1907, P. freudenreichii 169TM, P. freudenreichii ssp. freudenreichii 111, P. shermanii B-123, P. acidipropinici TL2 and P. acidipropionici P124I were able to pass all the tests and were considered as novel putative probiotic propionibacteria
Keywords: Dairy propionibacteria; probiotic; antimicrobial activity; bile salt
Microorganisms play a very important role in human health and nutrition. They are involved in the production of various bio-molecules, fermentation of milk and reside symbiotically in gut benefiting the host. Propionibacteria are one such important class of organisms. Propionibacteria are mesophilic, Gram positive, catalase positive, non-motile pleomorphic rods, non-sporeforming, and anaerobic to aerotolerant bacteria. Some cells may be elongated, bifid or arranged in "Chinese characters" [1]. The optimal growth o o temperature of these bacteria is between 30 C and 37 C [2].
Traditionally, propionibacteria are divided into two main groups: "the classical or dairy'' and "cutaneous" propioni bacteria. The dairy species includes Propionibacterium acidipropionici, P. jensenii, P. thoenii and both subspecies of P. freudenreichii. Propionibacteria play an important role in the production of flavor compounds and in the ripening of Swiss type cheese.
The literature concerning the potential probiotic properties of propionibacteria is very limited compared to that about lactobacilli and bifidobacteria. However, in recent years an increasing number of reports on the potential health benefits of propionbacteria have been published. The dairy propionibacteria have a number of properties that make them good probiotic candidates [3]. In this sense, P. acidipropionici P124I were able to pass all the tests and were Propionibacterium ssp. are able to produce a wide variety of biological compounds that enhance the human health like folic acid, proline, conjugated linoleic acid and vitamin B 12 [4], and synthesize several different bio-protective compounds such as bacteriocins or antifungal compounds [5]. Moreover, many members of the dairy group have a long history of safe use in food manufacturing [6].
Some strains of propionibacteria have a potential to be used as probiotic cultures. These strains produce bifidogenic compounds and show the ability to survive and maintain activity during passage of the digestive tract [2]. Furthermore, propionibacteria are able to stimulate the immune system and limit cancer progression although the mechanism involved is not defined. Cousin et al. [2] reported that dairy propionibacteria are able to prevent infections and allergies, promote immune system maturation, and reduce the risk of cancer because they bind carcinogenic compounds (mycotoxins, plants lactins and heavy metals). The search for strains which show resistance to biological barriers of the human gastrointestinal tract, and which possess physiological characteristics compatible with probiotic properties among propionibacteria, may eventually lead to the finding of new probiotic strains for functional food products. As part of the selection of new probiotic candidates, nine propionibacteria strains were subjected to a series of in vitro analysis to assess their probiotic properties.
Table (1) reports the species and strains of propionibacteria and indictor strains and their sources. To prepare active cultures, Propionibacterium strains were activated by their successive transfers every 48 h in yeast extract /lactate broth o (YELB) medium under anaerobic conditions at 30 C [7,8]. Stock cultures were prepared by maintaining strains in o YELB containing 30% glycerol at -80 C for further use. All indicator organisms were routinely prepared in Tryptone Soya Broth (Oxoid, Basingstoke, England) and grown at o 30 C for 24 h with exception of the mold strains, which were incubated for 72 h. The indicator strains were o maintained as frozen stock at -80 C in the presence of 150 ml/l glycerol as cryoprotective agent. All working cultures were prepared from stock cultures through subcultured twice before use in the manipulations.
Propionibacterium thoenii TL18 Institute National de la Propionibacterium Recherache Agronomique acidipropionici TL2 (INRA).
Propionibacterium American Type Culture freudenreichii ssp. Collection (ATCC), shermaniiATCC 1907 Manasses, USA.
Propionibacterium Bio.Vet.Inc.USA.
freudenreichii 169TM Propionibacterium University of Warmia and freudenreichii ssp. freudenreichii Mazury (Olsztn,Poland) 111 Propionibacterium shermanii B- Department of Food 123 Techn., propionibacteria culture collection, Iwa State Univ.
acidipropionici P124I Dairy Microbiology Bacillus subtililus Laboratory, National Bacillus cereus Research Center, Dokki, Escherichia coli Giza, Egypt.
Staphylococcus aureus Pseudomonas aeruginosa Saccharomyces cervisiae Candida albicans Alternaria sp Aspergillus niger Fusarium sp
o Cultures of propionibacteria were routinely grown at 30 C in YELB for 48 h under anaerobic conditions. After growing, morphological characteristics were determined with brightfield microscopy of Gram stained preparations, catalase activity and oxidase test, gas produced from glucose, growth in 20% bile salt, motility after 48 h in MRS medium [9] and colony pigmentation after 10 days on yeast extract /lactate agar (YELA) plates [8]. The strains were tested for production of acids from carbohydrates and related compounds by using API 50 CH strips (bio-Merieux, Fance). The tests were carried out according to the instructions of the manufacturer and results read after o incubation of strains at 30 C for 2, 4 and 7 days.
The experiment for tolerance of propionibacteria to pH 2, 3, 3.5 and 7(control) was performed following the method described by Sahadeva et al. [10] and Yuksedag et al. [8]. Viable cell counts were determined on YELA at various o times (0, 1, 2 and 3 h) of incubation at 37 C. The plates were o incubated at 30 C for 2 days under anaerobic conditions. Acid tolerance was estimated by comparing the growth of viable cell counts in all the YELA plates after 48 h.
To study the effect of bile salts on the growth of propionibacteria strains, a method described by Sahadeva et al. [10] was employed. All cultures were grown in YELB o for 24 h at 30 C. Bile resistance was determined on YELA containing different concentrations of oxal bile salts (Oxoid) (YEL-bile) (0, 0.1, 0.2, 0.3, 0.5 and 1% w/v). Viable counts were determined on YEL- bile agar and YELA (control) by plating serial dilutions in saline (0.9 % NaCl). Plates were o incubated for 48 h at 30 C anaerobically and only plates with 30 to 300 colonies were considered.
All propionibacteria strains were tested for antimicrobial activity against indicator strains using the agar diffusion method [9]. Test microorganisms were propagated twice and then grown for 18- 24 h. Propionibacteria species were inoculated in reconstituted skim milk powder (RSMP) supplemented with 1% yeast extract and 0.5% glucose (pH o 6.8) at 30 C for 8 days under anaerobic conditions. Activated cultures were centrifuged at 4000 rpm for 15 min and the clear supernatant was sterilized by filtration (0.45 μl), thus obtaining cell free filtrates.
6 An initial inoculum of approximately 10 cfu/ml of the target strain was incorporated into soft agar (1% w/v) plates of the appropriate for the target strain medium. Cell free filtrates (100 μl) were transferred in the holes (5mm diameter) drilled into the agar. The plates were incubated at o 30 C, depending on the target strain, and antimicrobial activity was recorded at growth free inhibition zones (diameter) around the well.
Carbohydrate fermentation profiles of the different strains were established using commercial API 50 CH system (bio Merieux, Marcyl' Etoile, France). Following the manufactures instructions [11].
Tolerance to phenol was investigated using YELA containing 0, 0.1, 0.2, 0.3, 0.4 and 0.5% phenol [12]. Plates o were incubated anaerobically at 30 C for 48 h and viable counts were determined.
A time-course experiment to study acid production profiles of Propionibacterium was carried out as described by Arici et al. [13]. The strains were grown in 100 ml of heat treated o (5 min at 121 C) 11% (w/v) reconstituted skim milk. Flasks o were removed after 18and 36 h of incubation at 30 C. The pH of fermented milk was measured directly by using a pH meter (Hanna, model 211).
Propionibacteria strains were screened for (EPS) production according to the method described by Mabrouk et al. [14]. Strains were grown on YELA medium supplemented with 100 g/l sucrose. The poured plates were incubated for 48 h o at 30 C under anaerobic conditions. At the end of incubation period, the bigger size and mucus colonies were tested for compactness or ropiness by touching them with a sterile metal loop and observed the formation of slime or strings.
The antibiotic susceptibility of propionibacteria strains were assessed using antibiotic discs diffusion method on MuellerHinton agar plates. Broth cultures of propionibacteria was 6 prepared using YELB and adjusted to 10 cfu/ml dilution of bacteria. A suspension of freshly grown Propionibacterium cultures of this concentration was spread on the surface of Mueller-Hinton agar plates. Two antibiotic discs (Oxoid) were placed on the surface of agar and the plates were o incubated at 30 C for 4 days, under anaerobic conditions. The susceptibility or resistance of the examined strains against the used antibiotics was assessed by measuring the diameter of the inhibition zone around the discs in mm. Strains were classified as resistant (-), moderately susceptible (+), susceptible (++), according to the recommendations published by the Clinical and Laboratory Standards Institute (2012) [15]. The Susceptibility pattern was assessed using Nalidixic acid (NA 30μg/ml), Tetracyclins (TET 30μg/ml), Trimethoprime/ Sulfamethoxazole (SXT 1.25/23.75μg), Polymyxin B (PB 30U/ml), Nitrofurantoin (F 300μg/ml), Clindamycin (CLi 2μg/ml), Imienem (Imi 10μg/ml), Aztreonam (Azt 30μg/ml), Norfloxacin (Nor 10μg), Cefazidime (Cef 30μg/ml), Gentamycin (CN 10μg/ml), PenicillinG (P 10u/ml), Ampicillin (Amp 10μg/ml), Chloroamphenicol (C 30μg/ml).
All the experiments were performed in triplicates and the data shown are the average of all repetitions.
Statistical analysis was carried out using Costat software program. Standard Error Mean (SEM) was carried out according to Fisher [16], LSD (List Significant Difference) test was used to compare the significant difference between means of treatment [17].
All the tested cultures were Gram positive in nature, catalase positive and short or filamentous rods in arrangements that resemble V, Y or Chinese characters. According to these morphological characteristics and as a result of the API identification tests all strains were grouped as dairy propionibacteria. Based on these results and the schemes for identifying species developed by Martinez et al. [18] and API 50 STREP, 9 strains were identified as 3 P. thoenii, 2 P. freudenreichii, 2 P. acidipropionici, and 1 for each P. shermanii and P. freudenreichii spp. shermanii. So this confirmed that all strains resembled the characteristics of genus Propionibacterium as described by Cummins and Johnson [19].
The ability of the Propionibacterium strains to tolerate acid is commonly used as one of the preliminary selection criteria for potential probiotic candidates. It was found that 8 Propionibacterium strains (89 %) out of 9 strains were able to retain viable counts above 6 log cfu/ml in pH 3 and 3.5 after 3 hours of incubation (Figure 1). Reduction in counts of between 0.4 and one logarithmic unit were commonly recorded for all strains, except P. thoenii P127, which showed more reduction in counts with a 1.5 and 2.1 -log reduction in viability at this pH. At pH 2.0, all tested strains showed progressive reduction in resistance during incubation, especially the strains of P. thoenii TL18, P. acidipropionici TL2, P. acidipropionici P124I, P. freudenreichii ssp. freudenreichii 111, P. freudenreichii 169TM, P. thoenii P15 and P. thoenii P127 which decreased by about 1.6 to 4 log units after 3 hours. Only two strains (P.freudenreichii ssp. shermanii (ATCC1907) and P. shermanii B-123) showed a high level of survival above 83% after 3 h of incubation (Figure 1).
From these results it is clear that more loss of viability of propionibacteria at pH 2.5 after 3 h could be observed. In this respect, Suoumalainen et al. [20] recorded that counts of propionibacteria was significantly reduced after 3h at pH 2
but not at pH 3 or 4.In the same line Darilmaz and Beyetli [21] found a significant reduction in cell count at pH 2 and 3 for P. freudenreichii ssp. freudenreichii and P. jensenii. On the other hand, Campaniello et al. [1] reported that at pH2.5 strains of propionibacteria did not show any kind of viability loss after 3 h. In the same point, Zarate et al. [22] mentioned
P.freudenreichii ssp. P.thoenii TL18
Bile tolerance is essential for probiotic strains to colonize in the small intestine [23]. Our strains of propionibacteria were suspended in sodium lactate agar media containing 0.1, 0.2, 0.3 and 0.5 % oxgall and without as control. Survival of o bacteria was determined after 48 hrs at 30 C. The statistical analysis of Table (2) showed that the concentrations of bile salts had not significant effects on the nine Propionibacterium strains tested.
As shown in Table (2) all tested strains showed a good survival at 0.1, 0.2, 0.3 and 0.5 % bile salt with some minor difference. The loss of log count reached 0.4, 0.5, 0.2, 0.5, 0.3, 0.4, 0.3, 0.7 and 0.3 log count in case of P. thoenii TL18, P. acidipropioniciTL2, P. acidipropionici P124I, P.
shermanii B-123, P. freudenreichii ssp. freudenreichii 111, P. freudenreichii 169TM, P. thoenii P15, P. freudenreichii ssp. shermanii (ATCC1907), P. thoenii P127, respectively.
On the other hand at concentration 1% , P.thoenii TL18, P.
that 4 Propionibacterium strains were shown to tolerate well at pH 4 and one strain lost viability at pH 3. While, all tested strains lost viability at pH 2. The difference between tested strains found could be due to the strong strain –specificity at acidic pH [1].
o
111
I P. freudenreichii169TM 111, P. thoenii P15 and P. thoenii P127 could not be detected after (48 hrs) this means a loss of viability. Other strains tested, P. acidipropionici P124I, P. freudenreichii 169TM, P. freudenreichii ssp. shermanii (ATCC1907) and P. thoenii P127 showed a good resistance as the loss of log -1 count reached 0.4, 0.6, 0.5 and 0.8 log cfu ml for these strains, respectively. From these results, it is clear that out of nine strains tested four strains showed high level of resistance while five strains have some resistance at concentration ranged from 0.1 to 0.5% of bile salt, but at 1.0 % strains P. thoenii TL18, P. acidipropioniciTL2, P. freudenreichii ssp. freudenreichii 111, P. thoenii P15, P. thoenii P127were inhibited completely. In this respect, Darilmaz and Beyatli [21] tested the resistance of some strains of propionibacteria to bile salt at concentrations 0.06, 0.15, 0.3 and 0.6%, they found that the tested strains were highly susceptible to these concentrations. In Contrary, Campeniello et al. [1] recorded that at 0.3% bile salt, ranging from 0.6 to 1.3 log cfu/ml after 24 hrs of incubation o at 30 C. In the same line Darilmaz and Beyatli [21] found that most of 29 strains of propionibacteria tested for their resistance to 0.6% bile salt had a high level of resistance.
But only seven strains showed some reduction in their resistance in the presence of 0.3% bile salt. Moreover, Yuksekdag et al. [8] recorded that different bile salt concentrations have a little effect on the tolerance of dairy propionibacteria, they recorded that bile salt did not seem to be related to the species but to the strain.
Naturally produced metabolic products of probiotic bacteria such as bacteriocins and organic acid are said to have inhibitory effect against some pathogenic microorganisms [24, 25]. Dairy propionibacteria produce antimicrobial agents that can inhibit the growth of spoilage and pathogenic microorganisms [26, 9, 24].
The statistical analysis of Table (3) showed a significant differences (P >0.05) were recorded among nine Propionibacterium strains toward various indicator pathogenic strains. .From Table (3) it is clear that all tested Propionibacterium have an inhibitory effect against B.
cereus but two strains only P. acidipropionici P124I and P.
freudenreichii 169 TM did not inhibit the growth of B.
subtilis. Only P. thoenii TL18 showed no inhibitory effect on E. coli and Pseudomonas aeruginosa. P.acidipropionici TL2 also didn't inhibit the growth of Staphylococcus aureus.
On the other hand, other tested Propionibacterium strains can inhibit the growth of these organisms. Saccharomyces cervisiae was inhibited by P.acidipropionici TL2, , P.
freudenreichii ssp. shermanii (ATCC1907), P.freudenreichii ssp. freudenreichii 111, P.thoenii P15, P.thoenii TL18 and P.
shermanii B -123but Candida albicans could be inhibited by P. thoenii P127, P. acidipropionici TL2, P. freudenreichii ssp. freudenreichii 111, P.shermanii B-123, P.freudenreichii ssp . shermanii (ATCC1907). Other propioni bacteria tested did not affect the growth of yeasts. Aspergillus niger and Alternaria are more resistant to inhibit by these bacteria as two strains P.thoenii P15 and P. shermanii B -123 affect the growth of A. niger.
However, three strains (P. acidipropionici TL2, P. freudenreichii ssp. shermanii (ATCC1907) and P. shermanii B-123) showed antimicrobial properties against Alternaria spp. Lastly, strains P. acidipropionici TL2, P. acidipropionici P124I, P. freudenrechii ssp. freudenrechii 111, P. freudenreichii ssp . shermanii (ATCC1907), P. shermanii B- 123 and P. thoenii P127 can inhibit the growth of Fusarium spp. It is clear that among 9 tested Propioni bacterium strains one strain P. shermanii B123 had a broad spectrum against pathogenic strains followed by P. freudenreichii ssp . shermanii (ATCC1907). The most sensitive organisms tested were B. cereus, followed by Pseudomonas aeruginosa and Staphylococcus aureus. Our results are in the same line of Yuksekdag et al. [8] as they recorded that antimicrobial effect of Propionibacterium ssp. strains was more obvious against the Gram negative strains. They added that S. aureus and B. cerues were sensitive to the metabolites of Propioni bacterium ssp. It is obvious that, there are differences in the degree of antagonistic effect depending on the indicator organism. It is worthy to mention that Darilmaz and Beyatli [21] reported that the different pathogens generally inhibited by propionibacteria.
Table (4) shows the strains ability to ferment various carbohydrates. All strains fermented glycerol, D-arabinose, galactose, glucose, fructose, mannose and sucrose. On the other hand, most strains showed positive results in the fermentation of the following sugars: ribose (eight of nine), melibiose (sex of nine), a-methyl D-mannoside and N- acethyl-glucosamine (five of nine). In addition, four strains fermented each of adonitol, sorbitol, lactose, rafinose, tagatose and gluconate. Sorbose fermented only with P. acidipropionici P124I, while turanose fermented with two strains (P. acidipropionici P124I and P. acidipropionici TL2). None of the nine propioni bacterium strains able to ferment a-methyl-D-glucoside and gentibiose. These results are similar to those obtained by Martinez et al. [18] when different potentially probiotic strains of propionibacteria were studied.
o -Values that have the same small letters in the same row are not significantly different (p>0.05).- n.d: not detected GLY: glycerol, ERY: erythritol, DAR:D-arabinose, RIB: ribose, ADO: adonitol, GAL: galactose, GLU: glucose, FRU: fructose, MNE: mannose, SBE: sorbose, INO:inositol, MAN: mannitol, SOR: sorbitol, MDM: a-methylD-mannoside,MDG: a-methyl-D-glucoside, NAG: N-acethyl-glucosamine, SAL: salicin, MAL: maltose, LAC: lactose, MEL:melibiose, SAC: sucrose, RAF: raffinose, GEN: gentibiose, TUR: turanose, TAG: tagatose, GNT: gluconate.
P.3: P.thoenii TL18 , P.6 : P.freudenreichii ssp. freudenreichii ssp 111,
Strains : P.1: P.thoenii P15, P.2:P.thoenii P127 , P.freudenreichii 169 TM , P.acidipropioniciP124I Symbols: +: present character, - : absence character
Resistance to phenol was tested as an additional indicator for survival under intestinal conditions for the same nine C. albicansS. S. aureus Fusarium spAlternaria sp INOMANSORMDMMDGNAGSALLACMELSACRAFGENTURTAGGNT P.4 : P.freudenreichii spp . shermanii (ATCC1907), P.5 :
P7: P.shermanii B-123 , P.8: PacidipropioniciTL2, P.9:
probiotic candidate strains. Phenol is an intermediate of putrefactive process in the intestinal tract that may be formed by bacterial determination of some aromatic amino
Values that have the same small letters in the same row are not significantly different (p>0.05).-n.d: not detected acids derived from dietary or endogenous produced proteins [15]. Table (5) shows the survival of Propionibacterium strains at different concentrations of phenol. These strains showed different degrees of sensitivity towards this compound. It can be seen that 4 and 6 strains were completely inhibited at 0.4 % and 0.5% phenol, respectively.
Statistical analysis proved that; phenol concentrations had significant effect (P>0.05) on the viability of all Propioni bacterium strains examined. In this respect, some authors suggest that a 0.4% concentration of phenol causes a bacteriostatic action in some microorganisms [27]. In contrast, 3 strains included P. freudenreichii ssp.
freudenreichii 111, P. shermanii B-123 and P. freudenreichii ssp. shermanii (ATCC1907) exhibited a good phenol tolerance.
The acidity developmentof tested Propionibacterium strains are presented in Table (6). The results showed that the tested nine strains displayed various behaviours in their acidification activity. The statistical analysis showed that no significant differences (P>0.05) were observed between the tested strains .The pH values ranged from 5.13 to 5.71 after o 18h. After 36h of incubation at 30 C the pH of milk inoculated with Propionibcterium was less than 5.40. In general, the results show that all the tested Propionibacterium strains acidified milk well. The acidification characteristy by propionibacteria strains is required for reducing fermentation time when using them in production of dairy products and reducing contamination by spoilage and/or pathogenic microorganisms. This result may be considered as a consequence of its obligatory homofermentative metabolism, the propionic fermentation leads to the production of short chain fatty acids- acetate and propionate [28].
-
The potential of propionibacteria to produce exopolysaccharide is displayed in Table (7). The results show that four strains P. acidipropionici TL2, P. acidipropionici P124I, P. shermanii B-123 and P. freudenreichii ssp. shermanii (ATCC1907) gave ropy colonies and were positive for production of exopolysaccharide. These results are similar to those reported by Gorret et al. [29] for Propionibacterium strains. Generally, the exopolysaccharide produced by probiotic strains is extracellularly secreted as microbial polysaccharide present on the surface of many probiotics. The capsular structure of exopolysaccharide has been found to protect strains from unfavorable environment. Furthermore, Ren et al. [30] reported that probiotic strains produced exopolysaccharides have an ability to exhibit acid and bile tolerance. In this study, the four strains produced exopolysaccharide exhibited acid and bile tolerance.
Because of the risk that any antibiotic resistance bacteria will spread to intestinal microbiota. The absence of antibiotic resistance for probiotic is an important criterion. Propionibacterium strains were tested for their sensitivity to antibiotics as this criterion is taken into account during the evaluation of probiotic strains. From Table (8) it is clear that all tested strains showed high sensitivity to Tetracyclins and Trimethoprime/Sulfamethoxazole. On the other hand, seven strains P. thoenii TL18, P. acidipropionici P124I, P. shermanii B-123, P. freudenreichii ssp. freudenreichii 111, P. freudenreichii 169TM, P. freudenreichii ssp . shermanii (ATCC1907), P. thoenii P127 could resist Ampicillin while strains P. thoenii TL18, P. thoenii P15, P. freudenreichii 169TM, P. freudenreichii ssp. freudenreichii 111 resist chloroamphenicol.
0 Nalidixic acid (NA 30μg/ml), Tetracyclins (TET 30μg/ml), Trimethoprime/Sulfamethoxazole(SXT 1.25/23.75μg), Polymyxin B (PB 30U/ml), Nitrofurantoin (F 300μg/ml), Clindamycin(CLi 2μg/ml), Imienem(Imi 10μg/ml), Aztreonam(Azt 30μg/ml), Norfloxacin(Nor 10μg), Cefazidime(Cef 30μg/ml), Streptomycin (S 10μg/ml), Kanamycin (K 30μg/ml), Gentamycin(CN 10μg/ml), PenicillinG(P 10u/ml),
-Values that have the same small letters in the same row are not significantly different (p>0.05)
Resistant (-), moderately susceptible (+), susceptible (++).
Ampicillin(Amp 10μg/ml), Chloroamphenicol(C 30μg/ml) From the same Table, it could be recorded that P. freudenreichii ssp .shermanii (ATCC1907) did not show any resistance to antibiotic tested, while P. acidipropionici P124I, can resist 7 of these antibiotics. In this respect, Suoumalainen et al. [20] recorded that P. freudenreichii ssp. shermanii (ATCC1907) have a high minimal inhibitory concentration value for aminoglycosides, streptomycin, gentamycin, kanamycin. It is worthy to mention that some authors reported that the resistance to antibiotic is not problem for these microorganisms [31].
(+) The strain produce EPS, (-) The strain not produce EPS o
In conclusion, the data present in this study reveals that eight of tested Propioni bacterium strains were found to possess desirable in vitro probiotic properties. These strains are good candidates for further investigation to elucidate the effect of commercially available oligosaccharides as prebiotics on their survival and activity.