Carbohydrate preference, acid tolerance and bile tolerance in five strains of Bifidobacterium

AIMS: To assess the suitability of bifidobacteria for inclusion in synbiotic products on the basis of carbohydrate preference, acid and bile tolerance. METHODS AND RESULTS: Five strains of Bifidobacterium were analysed for their carbohydrate preference from 12 substrates. Maximum growth rates were used to compare substrate preferences. Galacto-oligosaccharides and isomalto-oligosaccharides were well utilized by all the test species. Most bacteria tested could also utilize at least one type of fructan molecule. To determine transit tolerance of potentially probiotic bifidobacteria, acid and bile resistance was tested. A wide range acid resistance was found. Bile tolerance also varied. CONCLUSIONS: GOS and IMO were generally well utilized by the tested species. Other substrates were used to different degrees by the different species. Most bifidobacteria are poorly resistant to strongly acidic conditions with the exception of Bifidobacterium lactis Bb12. Bile tolerances were widely variable and it was shown that caution should be exercised when using colorimetric methods to assess bile tolerance. SIGNIFICANCE AND IMPACT OF STUDY: The study allows the comparison of the properties of bifidobacteria, allowing a cost effective screen for the best species for use in synbiotic products to allow better survival and efficacy.     

Prebiotics enhance survival and prolong the retention period of specific probiotic inocula in an in vivo murine model

AIM: To identify novel prebiotics that could be used to maintain persistence of three representative probiotic strains in vivo. METHODS AND RESULTS: Test mice were treated with prebiotics soybean oligosaccharide (SOS), fructooligosaccharide (FOS) or inulin, followed by probiotics Lactobacillus acidophilus LAFTI L10 (L10), Bifidobacterium lactis LAFTI B94 (B94) or Lactobacillus casei L26 LAFTI (L26). Faecal samples were then collected and analysed using selective medium and PCR analysis to determine the presence of the probiotic strains. In contrast to the control groups, in mice fed prebiotics, the survival and retention time of the test probiotics was increased extensively. SOS and FOS prolonged the retention period of L10 from 24 to 30 h. Of the three prebiotics, FOS gave the best result with B94, prolonging the retention period from 3 to > or =10 days. Of the three prebiotics, inulin gave the best result for L26, prolonging the retention period from 2 to > or =6 days. CONCLUSIONS: The prebiotics SOS, FOS and inulin significantly enhance survival and prolong the retention period of L10, B94 and L26 in vivo. SIGNIFICANCE AND IMPACT OF THE STUDY: Our results demonstrate the potential use of FOS, inulin and SOS as prebiotics in conjunction with the probiotic strains L10, B94 and L26 for new synbiotic products.     

Genetic heterogeneity and functional properties of intestinal bifidobacteria

AIMS: The aim of the present study was to compare several molecular methods for the identification and genotyping of bifidobacteria, and further to investigate genetic heterogeneity and functional properties of bifidobacterial isolates from intestinal samples of Finnish adult subjects. METHODS AND RESULTS: A total of 153 intestinal bifidobacterial isolates were included in initial screening and 34 isolates were further characterized. Identification results obtained with PCR-ELISA and ribotyping were well in accordance with each other, while randomly amplified polymorphic DNA (RAPD) gave tentative identification only to Bifidobacterium bifidum and to 65% of the B. longum isolates. The most commonly detected species were B. longum biotype longum followed by B. adolescentis and B. bifidum. In addition, B. animalis (lactis), B. angulatum and B. pseudocatenulatum were found. Ribotyping and pulsed-field gel electrophoresis (PFGE) proved to be discriminatory methods for bifidobacteria, but also RAPD revealed intraspecies heterogeneity. Besides two B. animalis (lactis) isolates with very close similarity to a commercially available probiotic strain, none of the intestinal isolates showed optimal survival in all tolerance (acid, bile and oxygen) or growth performance tests. CONCLUSIONS: Several species/strains of bifidobacteria simultaneously colonize the gastrointestinal tract of healthy Finnish adults and intestinal Bifidobacterium isolates were genetically heterogeneous. Functional properties of bifidobacteria were strain-dependent. SIGNIFICANCE AND IMPACT OF THE STUDY: Applicability of ribotyping with the automated RiboPrinter System for identification and genotyping of bifidobacteria was shown in the present study.     

Rapid identification of potentially probiotic Bifidobacterium species by multiplex PCR using species-specific primers based on the region extending from 16S rRNA through 23S rRNA

This study aimed at developing a novel multiplex polymerase chain reaction (PCR) primer set for identification of the potentially probiotic Bifidobacterium species B. adolescentis, B. animalis subsp. animalis (B. animalis), B. bifidum, B. breve, B. longum biovar infantis (B. infantis), B. animalis subsp. lactis B. lactis, B. longum biovar longum (B. longum) and B. pseudolongum. The primer set comprised specific and conserved primers and was derived from the integrated sequences of 16S and 23S rRNA genes and the rRNA intergenic spacer region (ISR) of each species. It could detect and identify type strains and isolates from pharmaceuticals or dairy products corresponding to the eight Bifidobacterium species with high specificity. It was also useful for screening of the related strains from natural sources such as the gastro-intestinal tract and feces. We suggest that the assay system from this study is an efficient tool for simple, rapid and reliable identification of Bifidobacterium species for which probiotic strains are known.     

Detection and quantification of Bifidobacterium lactis LAFTI B94 in human faecal samples from a consumption trial

A method was developed to allow detection of the probiotic Bifidobacterium lactis LAFTI B94 in human clinical samples. A new probe, Laf94p, was developed to accomplish colony hybridization of B. lactis B94. PCR detection of B94 was also achieved using the species-specific (B. lactis) primer pair. These tests and probes allowed detection and quantification of B94 in the human intestinal flora. The sensitivity of the probe was assessed by monitoring faecal levels of B94 in humans who were fed the culture. In this trial, five volunteers were fed with the probiotic. The presence of B94 was assessed daily. Viable B94 could be detected at high levels (as high as 1.8 x 10(9) cfu g(-1) wet weight) during the feeding period. Four weeks after the feeding stopped, B94 could still be detected in one subject. These results indicate that B94 survives in the human gastrointestinal tract.     

An in vitro model for investigating intestinal adhesion of potential dairy propionibacteria probiotic strains using cell line C2BBe1

AIMS: The purposes of this study were to screen the adhesion properties of dairy propionibacteria strains and evaluate whether C2BBe1 could be used in the screening of potential probiotic strains. METHODS AND RESULTS: Thirteen dairy propionibacteria strains and two control strains, Lactobacillus acidophilus MJLA1 and Bifidobacterium lactis BDBB2, were tested for adhesion to C2BBe1. Electron microscopic observations demonstrated that the control strains, L. acidophilus MJLA1 and B. lactis BDBB2, had similar adhesive ability to C2BBe1 as had been previously shown to Caco-2. Only one of the 13 strains of dairy propionibacteria, strain P. jensenii 702, demonstrated adhesion to C2BBe1. CONCLUSIONS: C2BBe1 can provide an alternative to Caco-2 for assessing in vitro adhesion properties of probiotic strains. Adhesion properties of dairy propionibacteria were strain-dependent. SIGNIFICANCE AND IMPACT OF THE STUDY: C2BBe1 is highly suitable for application in bacterial adhesion studies, and was used successfully to select a new potential probiotic.     

Employment of stressful conditions during culture production to enhance subsequent cold- and acid-tolerance of bifidobacteria

AIMS: This study examined whether exposure of early stationary phase Bifidobacterium longum and B. lactis cells to various combinations of reduced temperature, reduced pH and starvation would enhance the cells' subsequent cold- and/or acid-tolerance. METHODS AND RESULTS: Survival of B. longum in growth medium at 6 degrees C significantly (P < 0.05) increased as a result of starving cells for 30 or 60 min without any simultaneous decrease in temperature or pH. Acid-tolerance of B. lactis (at pH 3.5 in synthetic gastric fluid) increased significantly when the growth medium pH was decreased from 6.0 to 5.2 and cells experienced 30 or 60 min of starvation. Enhanced B. lactis acid-tolerance persisted through 8-11 weeks of -80 degrees C storage in the pH 5.2 growth medium. Upon addition to milk during yogurt manufacture, these cells initially had enhanced acid-tolerance relative to untreated cells but untreated cells became equally acid-tolerant during the first 2.5 h of yogurt manufacture. CONCLUSIONS: The cold- and acid-tolerance of bifidobacteria vary widely, but may be significantly increased by application of sub-lethal stress to early stationary phase cells during culture production. SIGNIFICANCE AND IMPACT OF THE STUDY: The enhancement of B. lactis acid-tolerance observed in this study may be of potential importance in the production of effective ready-to-consume probiotic dietary supplements.     

Increased stress tolerance of Bifidobacterium longum and Lactococcus lactis produced during continuous mixed-strain immobilized-cell fermentation

AIMS: The effect of immobilization and long-term continuous culture was studied on probiotic and technological characteristics of lactic acid and probiotic bacteria. METHODS AND RESULTS: A continuous culture in a two-stage system was carried out for 17 days at different temperatures ranging from 32 to 37 degrees C, with a first reactor containing Bifidobacterium longum ATCC 15707 and Lactococcus lactis subsp. lactis biovar. diacetylactis MD immobilized separately in gel beads, and a second reactor operated with free cells released from the first reactor. The tolerance of free cells from both strains produced in the effluent medium of both reactors to hydrogen peroxide, simulated gastric and intestinal juices, antibiotics and nisin, and freeze-drying markedly increased with culture time and was generally higher after 6 days than that of stationary-phase cells produced during free-cell batch fermentations. The reversibility of the acquired tolerance of B. longum, but not L. diacetylactis, to antibiotics was shown during successive free-cell batch cultures. CONCLUSIONS: Free cells produced from continuous immobilized-cell culture exhibited altered physiology and increased tolerance to various chemical and physico-chemical stresses. SIGNIFICANCE AND IMPACT OF THE STUDY: Continuous culture with immobilized cells could be used to produce probiotic and lactic acid bacteria with enhanced technological and probiotic characteristics.     

Specific identification and targeted characterization of Bifidobacterium lactis from different environmental isolates by a combined multiplex-PCR approach

The species Bifidobacterium lactis, with its main representative strain Bb12 (DSM 10140), is a yoghurt isolate used as a probiotic strain and is commercially applied in different types of yoghurts and infant formulas. In order to ensure the genetic identity and safety of this bacterial isolate, species- and strain-specific molecular tools for genetic fingerprinting must be available to identify isolated bifidobacteria or lactic acid bacteria from, e.g., various clinical environments of relevance in medical microbiology. Two opposing rRNA gene-targeted primers have been developed for specific detection of this microorganism by PCR. The specificity of this approach was evaluated and verified with DNA samples isolated from single and mixed cultures of bifidobacteria and lactobacilli (48 isolates, including the type strains of 29 Bifidobacterium and 9 Lactobacillus species). Furthermore, we performed a Multiplex-PCR using oligonucleotide primers targeting a specific region of the 16S rRNA gene for the genus Bifidobacterium and a conserved eubacterial 16S rDNA sequence. The specificity and sensitivity of this detection with a pure culture of B. lactis were, respectively, 100 bacteria/ml after 25 cycles of PCR and 1 to 10 bacteria/ml after a 50-cycle nested-PCR approach.     

Characterization of factors affecting attachment of Bifidobacterium species to amylomaize starch granules

AIMS: Two human-derived Bifidobacterium strains, PL1 and PL2, were tested for their ability to attach to amylomaize starch granules, and factors affecting binding were assessed. METHODS AND RESULTS: Good binding to granules was observed when the strains were grown on maltose or amylomaize starch, but not on glucose. Binding activity was localized to cell wall components and was sensitive to treatment with proteolytic enzymes. Several methodologies were employed to confirm these observations, including studies using radiolabelled cells, dot blot assays and scanning electron microscopy (SEM) analysis. CONCLUSION: Results from this study indicated that binding of strains PL1 and PL2 to amylomaize starch granules was mediated by a cell wall-associated proteinaceous factor that was induced when the strains were grown on starch or a related substrate, but not glucose. SIGNIFICANCE AND IMPACT OF THE STUDY: Attachment of probiotic strains to starch or other dietary fibres is believed to offer a selective advantage in the host intestine and may even prolong viability in adverse food environments. Therefore, characterizing the mechanisms of attachment has commercial implications in the design of synbiotic products.     

Predominant genera of fecal microbiota in children with atopic dermatitis are not altered by intake of probiotic bacteria Lactobacillus acidophilus NCFM and Bifidobacterium animalis subsp. lactis Bi-07

The effect of probiotic bacteria Lactobacillus acidophilus NCFM and Bifidobacterium lactis Bi-07 on the composition of the Lactobacillus group, Bifidobacterium and the total bacterial population in feces from young children with atopic dermatitis was investigated. The study included 50 children randomized to intake of one of the probiotic strain or placebo. Microbial composition was characterized by denaturing gradient gel electrophoresis, quantitative PCR and, in a subset of subjects, by pyrosequencing of the 16S rRNA gene. The core population of the Lactobacillus group was identified as Lactobacillus gasseri, Lactobacillus fermentum, Lactobacillus oris, Leuconostoc mesenteroides, while the bifidobacterial community included Bifidobacterium adolescentis, Bifidobacterium bifidum, Bifidobacterium longum and Bifidobacterium catenulatum. The fecal numbers of L. acidophilus and B. lactis increased significantly after intervention, indicating survival of the ingested bacteria. The levels of Bifidobacterium correlated positively (P=0.03), while the levels of the Lactobacillus group negatively (P=0.01) with improvement of atopic eczema evaluated by the Severity Scoring of Atopic Dermatitis index. This correlation was observed across the whole study cohort and not attributed to the probiotic intake. The main conclusion of the study is that administration of L. acidophilus NCFM and B. lactis Bi-07 does not affect the composition and diversity of the main bacterial populations in feces.     

A novel multi‐strain probiotic and synbiotic supplement for prevention of Clostridium difficile infection in a murine model

The protective effect of a multi‐strain probiotic and synbiotic formulation was evaluated in C57BL/6 mice infected with Clostridium difficile (CD) NAP1/027. Antibiotic‐treated mice were divided into the following four groups: Group 1, fed with a synbiotic formulation consisting of Lactobacillus plantarum F44, L. paracasei F8, Bifidobacterium breve 46, B. lactis 8:8, galacto‐oligosaccharides, isomalto‐oligosaccharides, and resistant starch; Group 2, fed with the same four probiotic strains as Group 1; Group 3, fed with the same prebiotic supplements as Group 1 for 7 days before CD infection; and Group 4 (control group) antibiotic treated and infected with NAP1/027 strain. Feces and cecal contents were collected for microbial cell viability, quantitative PCR (qPCR), toxin analyses and histopathology. Synbiotics‐ and probiotics‐fed mice showed a significant increase in total bifidobacteria (P < 0.05). The total lactobacilli count was increased in Group 1. Tests for cecal toxins were negative in Group 2 mice, whereas one sample each from Group 1 and 3 was positive. qPCR of cecal contents showed significant reduction in NAP1/027 DNA copies in Groups 1 and 2 and significantly higher numbers of B. breve 46, L. plantarum F44, and L. paracasei F8 in Groups 1 and 2 (P < 0.05); these changes were much less pronounced in Groups 3 and 4. Our findings indicate that the newly developed synbiotic or multi‐strain probiotic formulation confers protection against NAP1/027 infection in C57BL/6 mice. This holds promise for performing human studies.     

A comparative study of the preventative effects exerted by three probiotics, Bifidobacterium lactis, Lactobacillus casei and Lactobacillus acidophilus, in the TNBS model of rat colitis

AIMS: The intestinal anti-inflammatory effects of three probiotics with immunomodulatory properties, Lactobacillus casei, Lactobacillus acidophilus and Bifidobacterium lactis, were evaluated and compared in the trinitrobenzenesulphonic acid (TNBS) model of rat colitis. METHODS AND RESULTS: Colitis was induced in rats by intracolonic administration of 10 mg of TNBS dissolved in 0.25 ml of 50% ethanol. Each probiotic was administered orally (5x10(8) CFU suspended in 0.5 ml of skimmed milk) for 3 weeks, starting 2 weeks before the administration of TNBS. Colonic damage was evaluated histologically and biochemically 1 week after TNBS instillation. The results obtained revealed that all probiotics assayed showed intestinal anti-inflammatory effects, macroscopically evidenced by a significant reduction in the colonic weight/length ratio. Only B. lactis showed a lower incidence of diarrhoea in comparison with untreated rats. Biochemically, all probiotics restored colonic glutathione levels, depleted as a consequence of the oxidative stress of the inflammatory process. Bifidobacterium lactis treatment reduced colonic tumour necrosis factor (TNF)-alpha production, and inducible nitric oxide synthase (iNOS) and cyclo-oxygenase-2 (COX-2) expression; L. acidophilus administration reduced colonic leukotriene B4 production and iNOS expression and L. casei intake was associated with a decrease in colonic COX-2 expression. CONCLUSION: The three probiotics assayed have shown intestinal anti-inflammatory activity in the TNBS model of rat colitis, although each probiotic shows its own anti-inflammatory profile. SIGNIFICANCE AND IMPACT OF THE STUDY: These probiotics could be considered as potential adjuvants in the treatment of inflammatory bowel disease, although more studies are required in order to demonstrate their efficacy in humans.     

Effect of a synbiotic yogurt on levels of fecal bifidobacteria, clostridia, and enterobacteria

While ingestion of synbiotic yogurts containing Bifidobacterium animalis subsp. lactis and inulin is increasing, their effect on certain microbial groups in the human intestine is unclear. To further investigate this, a large-scale, crossover-design, placebo-controlled study was utilized to evaluate the effect of a synbiotic yogurt containing B. animalis subsp. lactis Bb-12 and inulin on the human intestinal bifidobacteria, clostridia, and enterobacteria. Fecal samples were collected at 14 time points from 46 volunteers who completed the study, and changes in the intestinal bacterial levels were monitored using real-time PCR. Strain Bb-12 could not be detected in feces after 2 weeks of washout. A live/dead PCR procedure indicated that the Bb-12 strain detected in the fecal samples was alive. A significant increase (P < 0.001) in the total bifidobacterial numbers was seen in both groups of subjects during the final washout period compared to the prefeeding period. This increase in total bifidobacteria corresponded with a significant decrease (P < 0.05) in numbers of clostridia but not enterobacteria. No significant differences in numbers of bifidobacteria, clostridia, or enterobacteria were observed between the probiotic and placebo groups during any of the feeding periods. However, subgrouping subjects based on lower initial bifidobacterial numbers or higher initial clostridial numbers did show corresponding significant differences between the synbiotic yogurt and placebo groups. This was not observed for a subgroup with higher initial enterobacterial numbers. While this synbiotic yogurt can increase bifidobacterial numbers and decrease clostridial numbers (but not enterobacterial numbers) in some individuals, it cannot modulate these microbial groups in the majority of individuals.     

Adhesion of bifidobacteria to granular starch and its implications in probiotic technologies

Adhesion of 19 Bifidobacterium strains to native maize, potato, oat, and barley starch granules was examined to investigate links between adhesion and substrate utilization and to determine if adhesion to starch could be exploited in probiotic food technologies. Starch adhesion was not characteristic of all the bifidobacteria tested. Adherent bacteria bound similarly to the different types of starch, and the binding capacity of the starch (number of bacteria per gram) correlated to the surface area of the granules. Highly adherent strains were able to hydrolyze the granular starches, but not all amylolytic strains were adherent, indicating that starch adhesion is not a prerequisite for efficient substrate utilization for all bifidobacteria. Adhesion was mediated by a cell surface protein(s). For the model organisms tested (Bifidobacterium adolescentis VTT E-001561 and Bifidobacterium pseudolongum ATCC 25526), adhesion appeared to be specific for alpha-1,4-linked glucose sugars, since adhesion was inhibited by maltose, maltodextrin, amylose, and soluble starch but not by trehalose, cellobiose, or lactose. In an in vitro gastric model, adhesion was inhibited both by the action of protease and at pH values of < or =3. Adhesion was not affected by bile, but the binding capacity of the starch was reduced by exposure to pancreatin. It may be possible to exploit adhesion of probiotic bifidobacteria to starch granules in microencapsulation technology and for synbiotic food applications.     

PCR-ELISA II: Analysis of Bifidobacterium populations in human faecal samples from a consumption trial with Bifidobacterium lactis Bb-12 and a galacto-oligosaccharide preparation

A PCR-ELISA method was extended for detection of most common Bifidobacterium species in humans and applied to a feeding trial including administration of Bifidobacterium lactis Bb-12 and galacto-oligosaccharide (GOS)-containing syrup as probiotic and prebiotic preparations, respectively. For PCR-ELISA, oligonucleotide probes based on 16S rDNA sequences were designed and tested for specificity and sensitivity with nine different bifidobacterial species followed by analysis of faecal samples. Bifidobacteria were monitored for their fluctuations during and after the feeding trial. Bifidobacterium longum was the most common species found in the faecal samples, followed by B. adolescentis and B. bifidum. During ingestion of the probiotic B. lactis Bb-12, the strain appeared in the faeces but was absent again one week after finishing of the trial. The species that were observed in the faecal samples taken prior to the feeding experiments persisted also in samples derived from the pre-feeding and feeding periods. The most consistent change observed was the decrease in the relative amount of B. longum in the test group ingesting either B. lactis Bb-12 alone or in combination with GOS-syrup. Since the amounts of B. longum increased again in the post-feeding sample with these subjects, it may suggest that to some extent B. lactis Bb-12 is able to transiently replace B. longum.     

Functional petit-suisse cheese: measure of the prebiotic effect

Prebiotics and probiotics are increasingly being used to produce potentially synbiotic foods, particularly through dairy products as vehicles. It is well known that both ingredients may offer benefits to improve the host health. This research aimed to evaluate the prebiotic potential of novel petit-suisse cheeses using an in vitro fermentation model. Five petit-suisse cheese formulations combining candidate prebiotics (inulin, oligofructose, honey) and probiotics (Lactobacillus acidophilus, Bifidobacterium lactis) were tested in vitro using sterile, stirred, batch culture fermentations with human faecal slurry. Measurement of prebiotic effect (MPE) values were generated comparing bacterial changes through determination of maximum growth rates of groups, rate of substrate assimilation and production of lactate and short chain fatty acids. Fastest fermentation and high lactic acid production, promoting increased growth rates of bifidobacteria and lactobacilli, were achieved with addition of prebiotics to a probiotic cheese (made using starter+probiotics). Addition of probiotic strains to control cheese (made using just a starter culture) also resulted in high lactic acid production. Highest MPE values were obtained with addition of prebiotics to a probiotic cheese, followed by addition of prebiotics and/or probiotics to a control cheese. Under the in vitro conditions used, cheese made with the combination of different prebiotics and probiotics resulted in the most promising functional petit-suisse cheese. The study allowed comparison of potentially functional petit-suisse cheeses and screening of preferred synbiotic potential for future market use.     

Growth kinetics on oligo- and polysaccharides and promising features of three antioxidative potential probiotic strains

Aims: To determine the antioxidative activity, glutathione production, acid and bile tolerance and carbohydrate preferences of Lactobacillus plantarum LP 1, Streptococcus thermophilus Z 57 and Bifidobacterium lactis B 933. Methods and Results: The intact bacteria exhibited antioxidative capacity against linolenic acid and ascorbate oxidation. The antioxidative activity of cell-free extracts was determined by chemiluminescent assay and agreed with total glutathione content. Superoxide dismutase was negligible in all the strains. Bile and gastric juice resistance was tested in vitro to estimate the transit tolerance in the upper gastrointestinal tract. Bifidobacterium lactis B 933 and L. plantarum LP 1 were more acid tolerant than S. thermophilus Z 57. All the strains were resistant to bile. Among 13 indigestible carbohydrates, galacto-oligosaccharides and fructo-oligosaccharides were utilized by all the strains and did not affect survival in human gastric juice. Conclusions: These potential probiotic strains exhibited antioxidative properties and good viability in gastric juice and bile may indicate tolerance to the transit through the upper gastrointestinal tract. Galacto-oligosaccharides and fructo-oligosaccharides are the most appropriate prebiotics to be used in effective synbiotic formulations. Significance and Impact of the Study: These results outline promising strains with antioxidative properties. Carbohydrate preferences can be exploited in order to develop synbiotic products.     

Evaluation of microencapsulation of a Bifidobacterium strain with starch as an approach to prolonging viability during storage

AIMS: To optimize a spray coating process for the production of encapsulated microspheres containing viable Bifidobacterium cells and to determine whether the readily gelatinized modified starch coating used in this study improved bacterial survival in foods or under acid conditions. METHODS AND RESULTS: An air inlet temperature of 100 degrees C was demonstrated to be optimal for the spray drying process, as it afforded good drying, low outlet temperatures (45 degrees C) and resulted in less than 1 log reduction in bifidobacteria numbers during drying. Maximum recovery yields of 30% were obtained after optimizing the air aspiration conditions. The average size of the Bifidobacterium PL1-containing starch microparticles was determined by scanning electron microscopy to be of the order of 5 microm. The starch-coated cells did not display any enhanced viability compared with free PL1 cells when exposed to acid conditions for 6 h or in two dry food preparations over 20 d storage at ambient temperature (19-24 degrees C). Determination of 1491 nucleotides of the 16S rRNA gene from PL1 indicated that it shared 97% homology with a previously sequenced Bifidobacterium ruminantium strain. CONCLUSIONS: Our data demonstrated that, although spray drying is a valuable process for encapsulating bifidobacteria, further work is required to ascertain a more appropriate coating material that will protect this strain against adverse environmental conditions. SIGNIFICANCE AND IMPACT OF THE STUDY: The production of small, uniformly coated microspheres containing viable bifidobacteria using an affordable and industrially convenient process, such as spray drying, has commercial implications for the production of probiotic products. Although popular for use as a coating polymer by the food industry, this study indicated that modified starches might not be suitable for use as an encapsulating material for probiotic strains.