Fermentum Strains Research Articles

Diversity of lactic acid bacteria from Hussuwa, a traditional African fermented sorghum food

The diversity of lactic acid bacteria associated with Hussuwa fermentation, a Sudanese fermented sorghum food, was studied using a polyphasic taxonomical approach. Predominant strains could be well characterised based on a combination of phenotypic tests and genotypic methods such as ARDRA, rep-PCR and RAPD-PCR, as well as 16S rRNA gene sequencing of representative strains. Thus, the majority (128 of 220, 58.3%) of strains exhibited phenotypic properties typical of heterofermentative lactobacilli and of these, 100 strains were characterised more closely using the genotyping methods. The majority (97/100) strains could be characterised as Lactobacillus fermentum strains. Seventy-two of 220 strains (32.7%) showed phenotypic properties that are characteristic of pediococci. Of 41 selected strains investigated by genotyping techniques, 38 (92.7%) could be characterised as Pediococcus acidilactici strains, while three (7.3%) could be characterised as Pediococcus pentosaceus strains. The Hussuwa fermentation thus appears to be dominated by L. fermentum strains and P. acidilactici strains. For this reason, we selected representative and predominant strains as potential starter cultures for Hussuwa fermentation. These strains, L. fermentum strains BFE 2442 and BFE 2282 and P. acidilactici strain BFE 2300, were shown on the basis of RAPD-PCR fingerprinting to predominate in a model fermentation when used as starter cultures inoculated at 1 × 10 6 CFU/g and to lower the pH of the fermentation to below pH 4.0 within 48 h. These cultures should be studied for further development as starter preparations in pilot scale studies in actual field fermentations.

Antimicrobial Activity, Inhibition of Urogenital Pathogens, and Synergistic Interactions Between Lactobacillus Strains

Lactobacillus fermentum strain L23 and L. rhamnosus strain L60 were selected as an alternative treatment to prevent or treat urogenital infections based on their probiotic properties and production of bacteriocins. The objectives of the present work were to study the inhibitory activities of these two bacteriocin-producing strains, and to analyze the interactions between pairs of bacteriocins that inhibit urogenital pathogens. Antimicrobial activity tests of L23 and L60 were performed by a diffusion method with 207 bacterial strains, isolated from female patients presenting a urogenital infection. Inhibitory substances interaction tests were carried out by using a streak-diffusion method on agar plates. One hundred percent of the clinical isolates showed sensitivity to the antimicrobial substances produced by L23 and L60. The selected lactobacilli produced larger inhibition halos when compared to several antibiotics commonly used for treating these infections. Synergistic interactions and indifferent interactions were recorded in 68.6% and 31.4% of the cases, respectively. No antagonistic interactions were observed. In conclusion, the bacteriocin-producing strains L23 and L60 are potential candidates for probiotic prophylaxis and treatment of urogenital disorders in women.

A Qualified Presumption of Safety approach for the safety assessment of Grana Padano whey starters

A Qualified Presumption of Safety (QPS) approach was applied to dominant lactic acid bacteria (LAB) associated with Grana Padano cheese whey starters. Thirty-two strains belonging to Lactobacillus helveticus, Lactobacillus delbrueckii subsp. lactis, Streptococcus thermophilus, and Lactobacillus fermentum, and representing the overall genotypic LAB diversity associated with 24 previously collected whey starters [Rossetti, L., Fornasari, M.E., Gatti, M., Lazzi, C., Neviani, E., Giraffa, G., 2008. Grana Padano cheese whey starters: microbial composition and strain distribution. International Journal of Food Microbiology 127, 168–171], were analyzed. All L. helveticus, L. delbrueckii subsp. lactis, and S. thermophilus isolates were susceptible to four (i.e. vancomycin, gentamicin, tetracycline, and erythromycin) of the clinically most relevant antibiotics. One L. fermentum strain displayed phenotypic resistance to tetracycline (Tet R), with MIC of 32 µg/ml, and gentamycin (Gm R), with MIC of 32 µg/ml. PCR was applied to this strain to test the presence of genes tet(L), tet(M), tet(S), and aac(6' )-aph(2' )-Ia, which are involved in horizontal transfer of Tet R and Gm R, respectively but no detectable amplification products were observed. According to QPS criteria, we conclude that Grana cheese whey starters do not present particular safety concerns.

Lactobacillus fermentum ME-3 - an antimicrobial and antioxidative probiotic.

The paper lays out the short scientific history and characteristics of the new probiotic Lactobacillus fermentum strain ME-3 DSM-14241, elaborated according to the regulations of WHO/FAO (2002). L. fermentum ME-3 is a unique strain of Lactobacillus species, having at the same time the antimicrobial and physiologically effective antioxidative properties and expressing health-promoting characteristics if consumed. Tartu University has patented this strain in Estonia (priority June 2001, patent in 2006), Russia (patent in 2006) and the USA (patent in 2007). The paper describes the process of the identification and molecular typing of this probiotic strain of human origin, its deposition in an international culture collection, and its safety assessment by laboratory tests and testing on experimental animals and volunteers. It has been established that L. fermentum strain ME-3 has double functional properties: antimicrobial activity against intestinal pathogens and high total antioxidative activity (TAA) and total antioxidative status (TAS) of intact cells and lysates, and it is characterized by a complete glutathione system: synthesis, uptake and redox turnover. The functional efficacy of the antimicrobial and antioxidative probiotic has been proven by the eradication of salmonellas and the reduction of liver and spleen granulomas in Salmonella Typhimurium-infected mice treated with the combination of ofloxacin and L. fermentum strain ME-3. Using capsules or foodstuffs enriched with L. fermentum ME-3, different clinical study designs (including double-blind, placebo-controlled, crossover studies) and different subjects (healthy volunteers, allergic patients and those recovering from a stroke), it has been shown that this probiotic increased the antioxidative activity of sera and improved the composition of the low-density lipid particles (LDL) and post-prandial lipids as well as oxidative stress status, thus demonstrating a remarkable anti-atherogenic effect. The elaboration of the probiotic L. fermentum strain ME-3 has drawn on wide international cooperative research and has taken more than 12 years altogether. The new ME-3 probiotic-containing products have been successfully marketed and sold in Baltic countries and Finland.

Enzyme activities of lactic acid bacteria from a pearl millet fermented gruel ( ben-saalga) of functional interest in nutrition

Lactic acid bacteria responsible for the fermentation of a pearl-millet based fermented gruel, ben-saalga, were investigated for enzyme activity in relation with the nutritional characteristics of gruels used as complementary foods for young children. Thirty pre-selected LAB from a set of 155 isolates were characterized principally for their ability to produce amylase, phytase and α-galactosidase. Two Lactobacillus plantarum strains (4.4 and 6.1) and three Lactobacillus fermentum strains (11.11.2, 3.7, 7.4) able to produce one or more of these enzymes were selected. Only weak amylase activity was found in the two Lactobacillus plantarum strains. α-amylase activity was associated with cells and was lower than 0.05 Ceralpha Units/ml. Phytase activity was detected in all five strains and was linked to the cell. The highest phytase activity was found in Lb. plantarum 4.4 and 6.1 (348.7 ± 17.4U/ml and 276.3 ± 51.4U/ml, respectively) and Lb. fermentum 7.4. (276.3 ± 13.2U/ml). All strains displayed a cell-linked α-galactosidase activity. In a medium containing 2% glucose, the highest cellular activity was found in Lb. fermentum 3.7 (1441.1 ± 133.7U/ml) and Lb. plantarum 4.4 (1223.1 ± 148.3U/ml) after 6h of fermentation in the presence of stachyose, and in Lb. plantarum 4.4 (763.3 ± 23.5U/ml) and Lb. fermentum 7.4 (346.7 ± 14.8U/ml) after 24h of fermentation with raffinose. These results are consistent with previous observations showing that phytates and α-galactooligosaccharides decreased during the natural lactic acid fermentation of pearl millet slurries, and that partial starch hydrolysis can be performed by endogenous microflora provided a pre-gelatinisation step is included in the process.

Functional characteristics of Lactobacillus spp. from traditional Maasai fermented milk products in Kenya

In this study functional characteristics of 23 representative Lactobacillus strains isolated from the Maasai traditional fermented milk ‘ Kule naoto’ were determined. The Lb. acidophilus group strains showed resistance to gastric juice and bile. In addition, some Lb. acidophilus strains expressed bile salt hydrolase activity, and had ability to assimilate cholesterol in vitro. In-vitro adhesion to HT29 MTX cells of up to 70% was recorded. Lb. fermentum strains showed almost 100% survival under simulated stomach acidic conditions and physiological salt concentrations of bile salts, hydrophobicity values were over 80%. Most strains of the Lb. casei and Lb. acidophilus groups showed aggregation abilities of above 50%. Many strains expressed a protective effect against N-methyl- N′-nitro— N-nitrosoguanidine induced DNA damage according to the ‘comet assay’ and none was virulent. The antibiotic minimum inhibitory concentration of selected strains was established. According to these results, the Lactobacillus spp associated with ‘ Kule naoto’, contain potentially probiotic (functional) strains.

Purification and Partial Characterization of Novel Bacteriocin L23 Produced by Lactobacillus fermentum L23

Lactobacillus fermentum strain L23 produced a small bacteriocin, designated bacteriocin L23, with an estimated molecular mass of < 7000 Da. Isolation, purification, and partial characterization of bacteriocin L23 are described. It displayed a wide inhibitory spectrum including both Gram-negative and Gram-positive pathogenic strains and two species of Candida. The antibacterial activity of cell-free culture supernatant fluid was not affected by catalase or urease but was abolished by the proteolytic enzymes trypsin and protease VI. Bacteriocin L23 was heat stable (60 min at 100 degrees C) and showed inhibitory activity over a wide pH range (4.0 to 7.0). The proteinaceous compound was isolated from cell-free culture supernatant fluid and purified. Crude bacteriocin sample was prepared by a process of ammonium sulfate precipitation, gel filtration, thin-layer chromatography, bioautography, and reversed-phase HPLC.

Rapid Detection and Quantification of Tyrosine Decarboxylase Gene (tdc) and Its Expression in Gram-Positive Bacteria Associated with Fermented Foods Using PCR-Based Methods

In this study, PCR-based procedures were developed to detect the occurrence and quantify the expression of the tyrosine decarboxylase gene (tdc) in gram-positive bacteria associated with fermented foods. Consensus primers were used in conventional and reverse transcription PCR to analyze a collection of 87 pure cultures of lactic acid bacteria and staphylococci. All enterococci, Staphylococcus epidermidis, Lactobacillus brevis, Lactobacillus curvatus, and Lactobacillus fermentum strains and1of 10 Staphylococcus xylosus strains produced amplification products with the primers DEC5 and DEC3 in accordance with results of the screening plate method and with previously reported result obtained with high-performance liquid chromatography. No amplicons were obtained for tyramine-negative strains, confirming the high specificity of these new primers. A novel quantitative real-time PCR assay was successfully applied to quantify tdc and its transcript in pure cultures and in meat and meat products. This assay allowed estimation of the influence of different variables (pH, temperature, and NaCl concentration) on the tdc expression of the tyraminogenic strain Enterococcus faecalis EF37 after 72 h of growth in M17 medium. Data obtained suggest that stressful conditions could induce greater tyrosine decarboxylase activity. The culture-independent PCR procedures developed here may be used for reliable and fast detection and quantification of bacterial tyraminogenic activity without the limitations of conventional techniques.

Different probiotic properties for Lactobacillus fermentum strains isolated from swine and poultry

Systematic procedures were used to evaluate the probiotic properties of Lactobacillus fermentum ( L. fermentum) strains isolated from swine and poultry. The major properties included their capabilities to adhere to the intestinal epithelium of swine and poultry, the inhibition on pathogenic bacteria, and their tolerance to the gastric juice and bile salts. Results showed that L. fermentum strains from poultry digestive tract showed better adherence to the swine intestine and chicken crop epithelial cells as compared to those strains from the swine origin. In addition, six strains from poultry and one strain from swine showed adhesion specificity to their own intestinal epithelium. Four poultry isolates and one swine isolate were able to adhere to the epithelial cells from both swine and chicken. For gastric juice and bile tolerance, most of the strains isolated from swine or poultry were acid tolerant but less strains were bile intolerant. The spent culture supernatant (SCS) of these L. fermentum strains showed antagonistic effect against the indicator bacteria, such as Escherichia coli, Salmonella spp., Shigella sonnei and some enterotoxigenic Staphylococcus aureus. From the above studies, some L. fermentum strains isolated from poultry were found to have the probiotic properties required for use in animal feed supplement. This study suggested that poultry digestive tract may serve as potential source for the isolation of probiotic lactic acid bacteria.

Bactericidal activity of culture fluid components of Lactobacillus fermentum strain 90 TS-4 (21) clone 3, and their capacity to modulate adhesion of Candida Albicans yeast-like fungi to vaginal epithelial cells

Antagonistic activities of L. fermentum strain 90 TS-4 (21), L. casei ATCC 27216, and L. acidophilus ATCC 4356 and bactericidal activity of lactobacillus culture fluid towards E. coli strain K12, S. aureus, and S. epidermidis test cultures were studied. The bactericidal effect of L. fermentum strain 90 TS-4 (21) clone 3 culture fluid preparation (pH 6.0) on the test cultures was dose-dependent. Adhesion of C. albicans yeast-like fungi to vaginal epitheliocytes was more pronounced for strains isolated from women with asymptomatic infection than for strains isolated from women with manifest forms. L. fermentum strain 90 TS-4 (21) clone 3 culture fluid preparation modulated adhesion of yeast-like fungi only if the fungal strain was initially highly adherent.

Antibiotic Susceptibility Profiles of Lactobacillus reuteri and Lactobacillus fermentum

Lactobacillus reuteri and Lactobacillus fermentum, which are commonly used as food processing aids and probiotics, can potentially act as reservoirs of antibiotic resistance genes. Acquired resistance genes may be transferred via the food chain or in the gastrointestinal tract to pathogenic bacteria. Knowledge of the distributions of antibiotic MICs for a species is needed when using a phenotypic method to assess the presence of acquired resistance genes. In the present study, 56 L. reuteri and 56 L. fermentum strains that differed by source and spatial and temporal origin were assessed for antibiotic susceptibility using an Etest kit and a broth microdilution protocol. L. fermentum strains displayed a uniform distribution of MICs for all six antibiotics tested. L. reuteri strains had a bimodal distribution of MICs or a distribution with MICs above the test range for 7 of the 14 antibiotics tested. Genetic relatedness was observed among L. reuteri strains with high MICs for both ampicillin and tetracycline and among strains with high MICs for both erythromycin and clindamycin. Results obtained with the Etest and the broth microdilution method corresponded well with each other. Thus, further research may make it possible to define microbiological breakpoints for distinguishing between strains with and without acquired resistance genes.

Diversity and technological properties of predominant lactic acid bacteria from fermented cassava used for the preparation of Gari, a traditional African food

Traditional fermentation of cassava is dominated by a lactic acid bacteria (LAB) population. Fermentation is important for improving product flavour and aroma as well as safety, especially by reduction of its toxic cyanogenic glucosides. The production of Gari from cassava in Benin typically occurs on a household or small industrial scale, and consequently suffers from inconsistent product quality and may not always be safe for consumption. Therefore, the diversity of LAB from a typical cassava fermentation for the preparation of Gari, and their technologically relevant characteristics were investigated with a view towards selection of appropriate starter cultures. A total of 139 predominant strains isolated from fermenting cassava were identified using phenotypic tests and genotypic methods such as rep-PCR and RAPD-PCR. DNA–DNA hybridisation and sequencing of the 16S rRNA genes were done for selected strains. Lactobacillus plantarum was the most abundantly isolated species (54.6% of isolates), followed by Leuconostoc fallax (22.3%) and Lactobacillus fermentum (18.0%). Lactobacillus brevis, Leuconostoc pseudomesenteroides and Weissella paramesenteroides were sporadically isolated. The L. plantarum strains were shown to be better acid producers and capable of faster acid production than the L. fallax or L. fermentum strains. The incidence of β-glucosidase (linamarase) activity was also highest among strains of this species. Production of antagonistic substances such as H 2O 2 and bacteriocins, however, was more common among L. fallax and L. fermentum strains. Strains of all three species were capable of utilising the indigestible sugars raffinose and stachyose. Therefore, a starter culture containing a mixture of strains from all three species was recommended.

Enhancement in Ex Vivo Phagocytic Capacity of Peritoneal Leukocytes in Mice by Oral Delivery of Various Lactic-Acid-Producing Bacteria

Lactic-acid-producing bacteria (LABs) are known to have immunomodulating activity. In the current study, various LABs were tested for their immunity-enhancing activity, especially the phagocytic activity of leukocytes. Viable but not heat-killed cells of Weissella kimchii strain PL9001, Lactobacillus fermentum strain PL9005, and L. plantarum strain PL9011 significantly increased the ex vivo phagocytic capacity of mouse peritoneal leukocytes to ingest fluorescein isothiocyanate (FITC)-labeled Escherichia coli in a strain-dependent manner. Results of this and previous studies suggest these LABs as candidates for new probiotics. This is the first report of the enhancement of peritoneal leukocyte activity of these species.

A new probiotic cheese with antioxidative and antimicrobial activity.

The aim of our study was to develop an original probiotic cheese based on the Estonian open-texture, smear-ripened, semisoft cheese "Pikantne." Cheese was produced by two methods using cheese starter cultures (Probat 505) in combination with 0.04% of probiotic Lactobacillus fermentum strain ME-3 (10(9) cfu/mL) with high antimicrobial activity and antioxidative properties. The probiotic Lactobacillus was added into milk simultaneously with starter cultures (cheese A) and into drained curd (cheese B). After addition of probiotic L. fermentum ME-3, the cheese composition, flavor, and aroma were comparable to the control cheese (score values = 4.5, 4.2, and 3.7 for control cheese, cheese A, and cheese B, respectively). Cheese A, which had good sensory properties, was chosen for further testing of viability and probiotic properties. The probiotic strain was found to withstand the technological processing of cheese, surviving and sustaining moderate antimicrobial and high antioxidative activity throughout ripening and storage (the ripened cheese contained approximately 5 x 10(7) cfu/g viable ME-3 cells), although the viability of the ME-3 strain incorporated into the cheese showed a slight decrease between d 24 and 54 after cheese preparation. Semisoft cheese "Pikantne" serves as a suitable carrier of antimicrobial and antioxidative L. fermentum ME-3.

Microbial formation of nitrite-cured pigment, nitrosylmyoglobin, from metmyoglobin in model systems and smoked fermented sausages by Lactobacillus fermentum strains and a commercial starter culture

Two Lactobacillus fermentum strains (JCM1173 and IFO3956) were evaluated for their ability to generate nitrosylated derivatives of myoglobin either in broth media or fermented sausages. For comparison, a commercial starter culture was also included. All bacteria species investigated converted brown metmyoglobin into red myoglobin derivatives when incubated separately in broth, but only the two lactobacilli showed a signal for nitrosylmyoglobin as measured by electron spin resonance spectroscopy. In smoked sausages with added bacteria culture the highest amount of nitrosylmyoglobin was observed in the centre of sausage with added L. fermentum, but colour formation in sausages with 60 ppm of nitrite added was more pronounced. An outer peripheral zone of all fermented sausages contained levels of nitrosylmyoglobin comparable to nitrite-cured sausages. Nitrogenous gasses from smoke may, however, cause this zone to be formed. Depending on a further optimisation of the processing parameters, the bacteria's ability to generate NO could form the basis for production of cured meat products without the use of nitrite/nitrate.

Gastrointestinal microbial community shifts observed following oral administration of a Lactobacillus fermentum strain to mice

The indigenous gastrointestinal microbiota acts as an integral defense against the colonisation of orally introduced microbes. Whilst this can be important in host protection, some introduced species, including lactobacilli, can have a positive impact on existing microbial communities. The interaction of a candidate probiotic strain of Lactobacillus fermentum within the gastrointestinal tract was monitored in a mouse model and its effect on the indigenous microbiota observed. L. fermentum KLD was administered via oro-gastric doses to mice with both a specific pathogen-free (SPF) and an ampicillin-depleted gut microbiota, containing no detectable lactobacilli. Its persistence was monitored by detection in faecal homogenates using culturing methods and polymerase chain reaction with L. fermentum specific primers. Microbial population shifts were observed using denaturing gradient gel electrophoresis (DGGE). L. fermentum KLD was detected within the gastrointestinal tract of SPF mice for up to 36 h, and for greater than 11 days in the ampicillin-treated mice. The administration resulted in substantial changes within the host Lactobacillus levels, determined by DGGE of 16S rDNA from faecal samples. Denaturing gradient profiles, from faecal samples collected at a range of pre- and post-dose intervals of groups of 10 SPF mice, indicated that several other constituents of the gastrointestinal community also fluctuated following dosing. These included Bifidobacterium and Eubacterium, which increased following KLD administration. The indigenous microbiota affected the persistence of L. fermentum KLD and in SPF mice the administration of this strain induced significant shifts in the indigenous microbial community.

Gastrointestinal microbial community shifts observed following oral administration of a Lactobacillus fermentum strain to mice

The indigenous gastrointestinal microbiota acts as an integral defense against the colonisation of orally introduced microbes. Whilst this can be important in host protection, some introduced species, including lactobacilli, can have a positive impact on existing microbial communities. The interaction of a candidate probiotic strain of Lactobacillus fermentum within the gastrointestinal tract was monitored in a mouse model and its effect on the indigenous microbiota observed. L. fermentum KLD was administered via oro-gastric doses to mice with both a specific pathogen-free (SPF) and an ampicillin-depleted gut microbiota, containing no detectable lactobacilli. Its persistence was monitored by detection in faecal homogenates using culturing methods and polymerase chain reaction with L. fermentum specific primers. Microbial population shifts were observed using denaturing gradient gel electrophoresis (DGGE). L. fermentum KLD was detected within the gastrointestinal tract of SPF mice for up to 36 h, and for greater than 11 days in the ampicillin-treated mice. The administration resulted in substantial changes within the host Lactobacillus levels, determined by DGGE of 16S rDNA from faecal samples. Denaturing gradient profiles, from faecal samples collected at a range of pre- and post-dose intervals of groups of 10 SPF mice, indicated that several other constituents of the gastrointestinal community also fluctuated following dosing. These included Bifidobacterium and Eubacterium, which increased following KLD administration. The indigenous microbiota affected the persistence of L. fermentum KLD and in SPF mice the administration of this strain induced significant shifts in the indigenous microbial community.

Biofilm Formation and Contamination of Cheese by Nonstarter Lactic Acid Bacteria in The Dairy Environment

Defects in cheese, such as undesirable flavors, gas formation, or white surface haze from calcium lactate crystals, can result from growth of nonstarter lactic acid bacteria (NSLAB). The potential for biofilm formation by NSLAB during cheese manufacturing, the effect of cleaning and sanitizing on the biofilm, and bacterial growth and formation of defects during ripening of the contaminated cheese were studied. Stirred-curd Cheddar cheese was made in the presence of stainless steel chips containing biofilms of either of two strains of erythromycin-resistant NSLAB (Lactobacillus curvatus strain JBL2126 or Lactobacillus fermentum strain AWL4001). During ripening, the cheese was assayed for total lactic acid bacteria, numbers of NSLAB, and percentage of lactic acid isomers. Biofilms of L. curvatus formed during cheese making survived the cleaning process and persisted in a subsequent batch of cheese. The starter culture also survived the cleaning process. Additionally, L. curvatus biofilms present in the vat dislodged, grew to high numbers, and caused a calcium lactate white haze defect in cheese during ripening. On the other hand, biofilms of L. fermentum sloughed off during cheese making but could not compete with other NSLAB present in cheese during ripening.Pulsed-field gel electrophoresis results verified the presence of the two biofilm strains during cheese making and in the ripening cheese. Probable contamination sites in the plant for other NSLAB isolated in the cheese were identified, thus supporting the hypothesis that resident NSLAB biofilms are a viable source of contamination in the dairy environment.

Evaluation of nitric oxide production by lactobacilli.

Six strains of Lactobacillus fermentum and Lactobacillus plantarum were investigated for nitric oxide (NO) production. First, the potential presence of NO synthase was examined. None of the strains of L. fermentum and L. plantarum examined produced NO from L-arginine under aerobic conditions. Interestingly, all L. fermentum strains expressed strong L-arginine deiminase activity. All L. fermentum strains produced NO in MRS broth, but the NO was found to be chemically derived from nitrite, which was produced by L. fermentum from nitrate present in the medium. Indeed all L. fermentum strains express nitrate reductase under anaerobic conditions. Moreover, one strain, L. fermentum LF1, had nitrate reductase activity under aerobic conditions. It was also found that L. fermentum strains JCM1173 and LF1 possessed ammonifying nitrite reductase. The latter strain also had denitrifying nitrite reductase activity at neutral pH under both anaerobic and aerobic conditions. The LF1 strain is thus capable of biochemically converting nitrate to NO. NO and nitrite produced from nitrate by lactobacilli may constitute a potential antimicrobial mechanism. studied in a rat acute liver injury model (Adawi et al. 1997). The results indicate that Lactobacillus plantarum DSM 9842 may possess NOS (Adawi et al. 1997). However, NO production from L-arginine has not been investigated in pure cultures of L. plantarum. According to the results of a 15N enrichment experiment, traces of (NO2-+NO3-)-N (total oxidised nitrogen: TON), which seemed to be formed by the resting cells of Lactobacillus fermentum IFO3956, appeared to be derived from L-arginine (Morita et al. 1997). Therefore, it was suggested that L. fermentum may possess a NOS. However, NO produced from L-arginine was not directly measured and a NOS inhibitor test was not performed by Morita et al. (1997). It is known that L-arginine deiminase (ADI) in bacteria may convert L-arginine to NH4+ (Cunin et al. 1986), which may be further oxidised to TON via nitrification by bacteria. Therefore, 15N enrichment experiments could not definitely conclude that L. fermentum possess NOS to convert L-arginine directly to NO. In this study, six Lactobacillus strains belonging to L. plantarum and L. fermentum were measured for NO production in MRS broth. The metabolism of nitrate and L-arginine by the Lactobacillus cell suspensions was also studied. The possibility that NO and nitrite production by lactobacilli may be a potential probiotic trait is also discussed.

Selection of Lactobacillus fermentum strains able to durably colonize the digestive tract of mice harboring a complex human flora

Sixteen Lactobacillus fermentum strains were isolated from the fecal microflora of pigs. They were challenged in vivo, in gnotobiotic mice harboring a porcine flora devoid of lactobacilli. Some of the strains were able to colonize the digestive tract of the mice at high population levels, whilst other strains were eliminated. Chromosomal DNA restriction analysis performed on both colonizing (C-type) and non-colonizing (NC-type) strains were analyzed by pulsed-field gel electrophoresis. Strains LEMPL9 (C-type) and LEM1.16A (NC-type) exhibited a high degree of homology. When challenged in mice harboring a simplified anaerobic flora, LEMPL9 was able to colonize the digestive tract of the animals whereas LEM1.16A was again eliminated. Mutagenesis performed on LEM1.16A allowed the isolation of a mutant able to colonize the gut of mice harboring the simplified anaerobic flora or a complex human flora, suggesting that genes involved in colonization process might have been affected to enhance the bacterium's colonization ability.