Int J Syst Evol Microbiol, 2005 Jan, 55(Pt 1), 401 - 4 Lactobacillus delbrueckii subsp . indicus subsp . nov., isolated from Indian dairy products; Dellaglio F et al.; Four strains isolated from Indian dairy products and initially identified as Lactobacillus delbrueckii could not be assigned to a definite subspecies because molecular identification and phenotypic traits did not agree with those of recognized subspecies of L . delbrueckii . Hybridization of total DNA (78-86 % against type strains of the other three subspecies), AFLP and RAPD-PCR fingerprints, phylogenetic analysis based on 16S rRNA gene sequences and sequence analysis of two coding genes (recA and hsp60), together with phenotypic profiles, indicated that the four strains form a coherent cluster and represent a novel subspecies, for which the name Lactobacillus delbrueckii subsp . indicus subsp . nov . is proposed . The type strain is NCC725(T) (=LMG 22083(T)=DSM 15996(T)). Int J Syst Evol Microbiol, 2005 Jan, 55(Pt 1), 197 - 207 Polyphasic study of wine Lactobacillus strains: taxonomic implications; Rodas AM et al.; One hundred and seventy-eight lactobacilli isolated from wine were characterized by a polyphasic approach . Strains were phenotypically identified at genus and species level by classical tests including the analysis of cell morphology, homo/heterofermentative character, sugar fermentation patterns, growth at different temperatures and the optical nature of the isomer of lactic acid produced from glucose . Molecular techniques such as random amplification of polymorphic DNA (RAPD), amplified 16S rDNA restriction analysis (16S-ARDRA), PFGE-RFLP and ribotyping were used to characterize strains, and their potential for identification and/or typing was evaluated . The information obtained with these techniques was processed with the BioNumerics software in order to analyse relationships existing between isolated strains and various reference species of the genus . Then, taxonomic dendrograms were obtained, and this information allowed the proposal of molecular procedures suitable for the identification and typing of these wine micro-organisms . The techniques useful for both identification and typing were RAPD and ribotyping, while 16S-ARDRA was only useful for identification and PFGE-RFLP only for typing purposes . The wine strains were identified as Lactobacillus brevis (19 strains), Lactobacillus collinoides (2 strains), Lactobacillus hilgardii (71 strains), Lactobacillus paracasei (13 strains), Lactobacillus pentosus (2 strains), Lactobacillus plantarum (34 strains) and Lactobacillus mali (10 strains). Int J Syst Evol Microbiol, 2005 Jan, 55(Pt 1), 83 - 5 Lactobacillus satsumensis sp . nov., isolated from mashes of shochu, a traditional Japanese distilled spirit made from fermented rice and other starchy materials; Endo A et al.; Gram-positive, rod-shaped, motile lactic acid bacteria (strains NRIC 0603, NRIC 0604(T), NRIC 0605 and NRIC 0606) were isolated from shochu mashes using an enrichment culture approach . These strains clustered in the Lactobacillus casei-Pediococcus group and were closely related to Lactobacillus nagelii and Lactobacillus mali on the basis of 16S rRNA gene sequence similarity . Levels of DNA-DNA relatedness revealed genotypic separation of the four isolates from the above two species . The isolates are therefore considered to represent a novel species, for which the name Lactobacillus satsumensis is proposed . The type strain is NRIC 0604(T) (=JCM 12392(T)=DSM 16230(T)). Int J Syst Evol Microbiol, 2005 Jan, 55(Pt 1), 77 - 82 Lactobacillus gastricus sp . nov., Lactobacillus antri sp . nov., Lactobacillus kalixensis sp . nov . and Lactobacillus ultunensis sp . nov., isolated from human stomach mucosa; Roos S et al.; In an attempt to study the composition of the Lactobacillus flora from mucosa of human stomach, 16S rRNA gene sequences of 129 isolates were analysed . For 15 of these, the results differed significantly from known sequences, and additional tests were performed to determine whether these isolates represented as yet unrecognized species . Phenotypic and genetic characteristics revealed that these isolates represented four novel Lactobacillus species . Two belong to the Lactobacillus reuteri and the other two to the Lactobacillus delbrueckii subgroup of Lactobacillus . The names Lactobacillus gastricus sp . nov., Lactobacillus antri sp . nov., Lactobacillus kalixensis sp . nov . and Lactobacillus ultunensis sp . nov . are proposed, with the respective type strains Kx156A7(T) (=LMG 22113(T)=DSM 16045(T)=CCUG 48454(T)), Kx146A4(T) (=LMG 22111(T)=DSM 16041(T)=CCUG 48456(T)), Kx127A2(T) (=LMG 22115(T)=DSM 16043(T)=CCUG 48459(T)) and Kx146C1(T) (=LMG 22117(T)=DSM 16047(T)=CCUG 48460(T)). Int J Syst Evol Microbiol, 2005 Jan, 55(Pt 1), 35 - 40 Lactobacillus rossii sp . nov., isolated from wheat sourdough; Corsetti A et al.; Screening of sourdough lactic acid bacteria for bacteriocin production resulted in the isolation of a Gram-positive, catalase-negative, non-spore-forming, non-motile rod bacterium (strain CS1(T)) that could not be associated with any previously described species . Comparative 16S rRNA gene sequence analysis recognized strain CS1(T) as a distinct member of the genus Lactobacillus . By a species-specific PCR strategy, five additional strains previously isolated from sourdoughs were found to belong to the same species as strain CS1(T), as confirmed by 16S rRNA gene sequence analysis . The closest related species were Lactobacillus durianis, Lactobacillus malefermentans and Lactobacillus suebicus, with which strain CS1(T) shared 93 % sequence similarity . For a further characterization of strain CS1(T), physiological (growth temperature, CO(2) production, hydrolysis of arginine, isomeric type of lactate, sugar fermentation) and chemotaxonomic (G+C content and peptidoglycan structure) properties were determined . Phenotypic characterization showed that strain CS1(T) was a member of the obligately heterofermentative group of the genus Lactobacillus . The DNA G+C content was 44.6 mol% . The peptidoglycan was of the A3alpha (l-lys-l-ser-l-Ala(2)) type . Physiological, biochemical and genotypic data, as well as results of DNA-DNA hybridization of genomic DNA with one of the closest phylogenetic relatives, L . durianis (34.3 %), indicated that strain CS1(T) represents a novel species of the genus Lactobacillus for which the name Lactobacillus rossii sp . nov . is proposed . The type strain of this species is CS1(T) (=ATCC BAA-822(T)=DSM 15814(T)). Int J Syst Evol Microbiol, 2005 Jan, 55(Pt 1), 31 - 4 Lactobacillus suntoryeus sp . nov., isolated from malt whisky distilleries; Cachat E et al.; Eight strains of Lactobacillus with identical partial 16S rRNA gene sequences and similar randomly amplified polymorphic DNA patterns were isolated from fermentation samples from Japanese and Scottish malt whisky distilleries . Phylogenetic analysis of almost complete 16S rRNA gene sequences from three representative strains (two from Japan, one from Scotland) placed them in the genus Lactobacillus as members of the Lactobacillus acidophilus group . Lactobacillus helveticus and Lactobacillus gallinarum were the most closely related species, with 16S rRNA gene similarities of 99.3 and 98.1 %, respectively . A similar phylogeny was derived from partial sequences of elongation factor Tu (tuf) genes in which the alleles from the three distillery isolates were identical and shared 99.0 % similarity with L . helveticus and L . gallinarum tuf genes . S-layer (slp) gene sequences suggested different relationships among the strains and the distillery isolates no longer formed a monophyletic group . The alleles from the Japanese and Scottish strains shared only 54 % similarity . Chromosomal DNA from the distillery strains gave DNA-DNA hybridization values between 79 and 100 % but showed less than 43 and 22 % reassociation with L . helveticus and L . gallinarum DNA, respectively . The name Lactobacillus suntoryeus sp . nov . is proposed for this novel taxon; the type strain is strain SA(T) (=LMG 22464(T)=NCIMB 14005(T)). J Dairy Sci, 2005 Feb, 88(2), 527 - 33 Clinical Effects of Lactobacillus acidophilus Strain L-92 on Perennial Allergic Rhinitis: A Double-Blind, Placebo-Controlled Study; Ishida Y et al.; Studies in animals have suggested that lactic acid bacteria alleviate allergic diseases, however, little information is available on their clinical effect on allergy in humans . Thus, we examined the efficacy of orally administered Lactobacillus acidophilus strain L-92 (L-92) on perennial allergic rhinitis . In a randomized, double-blind, placebo-controlled clinical trial, 49 patients with perennial allergic rhinitis were randomized to receive either 100 mL of heat-treated fermented milk containing L-92 (n = 25) or acidified milk without lactic acid bacteria (placebo; n = 24) for 8 wk . The severity of symptoms was evaluated based on the changes in the scores of clinical symptoms . Oral administration of milk fermented with L-92 resulted in a statistically significant improvement of nasal symptom-medication scores . Ocular symptom-medication scores of patients in the L-92 intervention group tended to improve compared with those in the placebo group . In addition, clear decreases of the scores of swelling and color of the nasal mucosa were observed in the L-92 intervention group at 6 and 8 wk after the start of ingestion of fermented milk . There were no significant differences in serum antihouse dust mite immunoglobulin E levels nor in T helper type 1/T helper type 2 ratio between the 2 groups . These results suggest that oral administration of L-92 can alleviate the symptoms of perennial allergic rhinitis, however, statistically significant changes were not shown in blood parameters. J Microbiol, 2004 Dec, 42(4), 315 - 8 Probiotication of tomato juice by lactic Acid bacteria; Yoon KY et al.; This study was undertaken to determine the suitability of tomato juice as a raw material for production of probiotic juice by four lactic acid bacteria (Latobacillus acidophilus LA39, Lactobacillus plantarum C3, Lactobacillus casei A4, and Lactobacillus delbrueckii D7) . Tomato juice was inoculated with a 24-h-old culture and incubated at 30 degrees C . Changes in pH, acidity, sugar content, and viable cell counts during fermentation under controlled conditions were measured . The lactic acid cultures reduced the pH to 4.1 or below and increased the acidity to 0.65% or higher, and the viable cell counts (CFU) reached nearly 1.0 to 9.0x10(9)/ml after 72 h fermentation . The viable cell counts of the four lactic acid bacteria in the fermented tomato juice ranged from 10(6) to 10(8) CFU/ml after 4 weeks of cold storage at 4 degrees C . Probiotic tomato juice could serve as a health beverage for vegetarians or consumers who are allergic to dairy products. J Microbiol Methods, 2005 Mar, 60(3), 353 - 63 Modified electroporation protocol for Lactobacilli isolated from the chicken crop facilitates transformation and the use of a genetic tool; Mason CK et al.; Isolates of Lactobacillus spp . from a collection of potentially probiotic strains isolated from the crops of broiler chickens were found to be non-electrotransformable using published techniques . One strain of Lactobacillus salivarius was shown to develop electrocompetence when an overnight culture was incubated in fresh medium . The effect was enhanced if glycine was incorporated into the fresh growth medium . When these modifications were applied to a number of other crop isolates of Lactobacillus spp., electrocompetence could be detected in approximately half the strains tested . Two temperature sensitive plasmid vectors that had been used for the genetic modification of other lactic acid bacteria were introduced into a crop strain of Lb . salivarius . Both showed temperature sensitivity at 42 degrees C and above but were relatively stable at 37 degrees C . The genetic tool harbouring an IS element allowed the delivery of the plasmid to multiple independent sites in the host chromosome . Harnessing such genetic tools will facilitate the future genetic analysis of the host bacterium. Microbiol Res, 2004, 159(4), 339 - 46 Potential of selected lactic acid bacteria to produce food compatible antifungal metabolites; De Muynck C et al.; The aim of this study was to assess the potential of lactic acid bacteria to inhibit the outgrowth of some common food-spoiling fungi . Culture supernatants of 17 Lactic acid bacterial strains as well as of three commercial probiotic cultures were evaluated for antifungal activity using an agar-diffusion method . The method parameters were chosen in order to reveal compounds for potential use in food (bio)preservation . Thirteen strains showed antifungal activity of which five strains were very promising: Lactobacillus acidophilus LMG 9433, L . amylovorus DSM 20532, L . brevis LMG 6906, L . coryniformis subsp . coryniformis LMG 9196 and L . plantarum LMG 6907 . Four of these five strains were further examined; it was found that the produced antifungal metabolites were pH-dependent . The exact chemical nature of these substances has not been revealed yet. Oral Health Prev Dent, 2003, 1(2), 157 - 62 Influence of dental restorative materials on salivary Streptococcus mutans and lactobacilli in the primary dentition; Willershausen B et al.; PURPOSE: When restoring deciduous teeth with extensive lesions, preformed stainless steel crowns are increasingly used in addition to resin-based materials . The aim of the present clinical study was to examine the influence of composites and stainless steel crowns on the cariogenic bacteria S . mutans and Lactobacilli by means of a commercially available salivary test (CRT bacteria) in 100 children . MATERIALS AND METHODS: For 50 children (30 boys, 20 girls, mean age 5.7 +/- 2.4 years) only composites have been used as a restorative material (an average of 9 fillings per child, total number: n=450), which had been in situ for an average of 15.9 months at the time of the examination . In another group of 50 children (33 boys, 17 girls, mean age 5.9 +/- 2.5 years), stainless steel crowns and composites have been used for restorations (an average of 3.5 stainless steel crowns per child, total number: n=174, an average of 7 composite fillings per child, total number n=348), which had been in place for a mean period of 17.2 months . RESULTS: The salivary examination of the children with composite restorations showed a high proportion of patients (64%, n=32) with high numbers (> or = 10(5)) of S . mutans and Lactobacilli (54%, n=27) . Only 22% (n=11) of the children with additional stainless steel crown restorations were found to have high numbers (> or = 10(5)) of S . mutans and Lactobacilli (34%, n=17) . CONCLUSION: This study shows a potential positive inhibitory effect of stainless steel crown restorations as compared to composite fillings with respect to the oral bacterial colonization. Oral Health Prev Dent, 2003, 1(3), 185 - 94 A ten-year cross-sectional and follow-up study of salivary flow rates and mutans streptococci and lactobacillus counts in elderly Swedish individuals; Fure S; PURPOSE: The whole salivary flow rates and microbial conditions were examined in a 10-year follow-up and cross-sectional study in a random sample of elderly inhabitants of Gothenburg . MATERIAL AND METHODS: Of the 208 individuals examined at baseline, forty-nine per cent participated in the follow-up, 56, 37 and nine persons, respectively, in the age groups of 65, 75 and 85 years . In addition, a new random sample of 98 individuals aged 55 year was examined . RESULTS: The mean secretion rate decreased significantly with increasing age in terms of unstimulated and stimulated whole saliva . Of the 200 participants, 50% were taking medication, which could have hyposalivatory side-effects . Persons with a daily intake of > or =4 drugs had significantly lower unstimulated and stimulated secretion rates . Forty-five persons reported subjective dryness in the mouth . The mean saliva secretion rates among these persons were significantly lower and the number of drugs consumed significantly higher than in people with no such complaints . The overall salivary counts of lactobacilli and mutants streptococci increased with age . Higher counts of these bacteria were found in persons wearing removable dentures than in persons without dentures . CONCLUSION: The salivary and microbial conditions ought to be continuously monitored in old people, in order to identify those who need oral health promotive measures. Appl Environ Microbiol, 2005 Jan, 71(1), 297 - 302 Starch-Binding Domain Affects Catalysis in Two Lactobacillus {alpha}-Amylases; Rodriguez-Sanoja R et al.; A new starch-binding domain (SBD) was recently described in alpha-amylases from three lactobacilli (Lactobacillus amylovorus, Lactobacillus plantarum, and Lactobacillus manihotivorans) . Usually, the SBD is formed by 100 amino acids, but the SBD sequences of the mentioned lactobacillus alpha-amylases consist of almost 500 amino acids that are organized in tandem repeats . The three lactobacillus amylase genes share more than 98% sequence identity . In spite of this identity, the SBD structures seem to be quite different . To investigate whether the observed differences in the SBDs have an effect on the hydrolytic capability of the enzymes, a kinetic study of L . amylovorus and L . plantarum amylases was developed, with both enzymes acting on several starch sources in granular and gelatinized forms . Results showed that the amylolytic capacities of these enzymes are quite different; the L . amylovorus alpha-amylase is, on average, 10 times more efficient than the L . plantarum enzyme in hydrolyzing all the tested polymeric starches, with only a minor difference in the adsorption capacities. J Biotechnol, 2005 Feb 23, 115(4), 405 - 12 Epub 2004 Nov 11. High-pressure inactivation of Saccharomyces cerevisiae and Lactobacillus plantarum at subzero temperatures; Perrier-Cornet JM et al.; High hydrostatic pressure is a new technology in the food processing industry, and is used for cold pasteurization of food products . However, the pressure inactivation of food-borne microorganisms requires very high pressures (generally more than 400MPa) and long pressure holding times (5min or more) . Carrying out pressure processing at low temperatures without freezing can reduce these parameters, which presently limit the application of this technology, in keeping the quality of fresh raw product . The yeast, Saccharomyces cerevisiae and the bacterium, Lactobacillus plantarum were pressurized for 10min at temperatures between -20 and 25 degrees C and pressure between 100 and 350MPa . Pressurization at subzero temperatures without freezing significantly enhanced the effect of pressure . For example, at a pressure of 150MPa, the decrease in temperature from ambient to -20 degrees C allowed an increase in the pressure-induced inactivation from less than 1log up to 7-8log for each microorganism studied . However, for comparable inactivation levels, the kinetics of microorganism inactivation did not differ, which suggests identical inactivation mechanisms . Implications of water thermodynamical properties like compression, protein denaturation, as well as membrane phase transitions, are discussed. Reprod Nutr Dev, 2004 Sep-Oct, 44(5), 449 - 57 Effects of the administration of Lactobacilli on body growth and on the metabolic profile in growing Maltese goat kids; Chiofalo V et al.; The aim of this study was to evaluate the effect of some lactobacilli on body growth and on the metabolic-nutritional status in growing goat kids . Twenty growing Maltese goat kids (10 Control and 10 Bios) were studied . The animals of the Bios group received a concentrate including 1 g x kg(-1) of SEB Bovino (spray-dried), Akron S.r.l., Italy, with non bacterial components: gum arabic, soybean meal, silicate alum of magnesium, and with bacterial components: 10(11) cfu kg(-1) each of Lactobacillus acidophilus, Lactobacillus salivarius, Lactobacillus reuteri . Monthly, bio-metric and weight evaluations were carried out on each animal and individual blood samples were taken . The Bios group showed the highest body weight (Control 19 vs . Bios 23 kg P < 0.001), anamorphosis (Control 71 vs . Bios 78 P < 0.05) and body proportion (Control 35 vs . Bios 41 P < 0.001) indices; the lowest levels of Non Esterified Fatty Acids (Control 0.778 vs . Bios 0.403 mmol L(-1) P < 0.001), triglycerides (Control 0.21 vs . Bios 0.18 mmol L(-1) P < 0.05), urea (Control 8.83 vs . Bios 7.65 mmol L(-1) P < 0.05) and the highest levels of Alkaline Phosphatase (Control 270 vs . Bios 851 U L(-1) P < 0.01) and Creatine Kinase (Control 173 vs . Bios 285 U L(-1) P < 0.01) . The results testify to the better metabolic activity of the Bios group which achieved, at the end of the trial (7 months old), about 99% of the morphological development of the adult, therefore an adequate structure for mating and going into production within the first year of life. Reprod Nutr Dev, 2004 Sep-Oct, 44(5), 397 - 405 Preliminary assays for the development of a probiotic for goats; Draksler D et al.; In order to determine probiotic properties, 137 strains of lactic acid bacteria from the feces of Creole goats were screened, only six were resistant to pH 2.0 and bile salts (0.3%) . Three strains identified as Lactobacillus and two as Enterococcus showed agglutination with the treated yeast . Between them, Lactobacillus DDL17, DDL19, DDL48 and Enterococcus DDE39 demonstrated high specificity in this test because the correspondent agglutination was inhibited by one sugar, suggesting the presence of a lectin-like structure in their cell walls, which could be due to adhesion ability . Another Enterococcus strain (DDE55) showed low affinity because five sugars inhibited the agglutination of the treated yeasts . The results of hydrophobic properties showed that the strains who were able to agglutinate yeasts presented similar hydrophobic characteristics as hexadecane, xylene and toluene, but high specificity was not related to a high hydrophobicity . Only two strains (Lactobacillus DDL19 and DDL48) showed aggregation with the lowest concentration of ammonium sulfate, complementing the hydrophobicity assay . Only one strain, Lactobacillus DDL48, showed an inhibition against an enteric indicator strain (Salmonella Typhimurium and Escherichia coli O111) . This inhibitory action was not affected by the addition of catalase and no inhibition was detected after neutralizing the supernatant culture fluid . These strains could be pre-selected in order to complete studies focused on designing a probiotic for use in goat feed. Zh Mikrobiol Epidemiol Immunobiol, 2004 Nov-Dec, (6), 101 - 6 {The role of cytokines in the immunoreactivity modulation with bacteria of the Lactobacillus genus} {Modulation of immune system cells by lactobacilli} {No authors listed} The influence of lactobacilli on the proliferative potential of immune system cells after the intragastral administration of viable microbial cells and their native filtrates to mice CBA is evaluated . The data have been obtained on the modulating influence of lactobacilli on the formation of T- and B-cell immune response--their role in maintaining homeostatsis and specific features of cell mediated immune reactions after the intragastral administration of virulent Shigella dysenteriae for modeling experimental infection in CBA mice . The mechanisms of the immunomodulating action of lactobacilli on local and systemic reactions of the host as well as realization of the protective properties of lactobacilli against the causative agents of acute enteric infections are discussed. Appl Microbiol Biotechnol . 2005 Jan 6; {Epub ahead of print} Cell surface display system for Lactococcus lactis: a novel development for oral vaccine; Raha AR et al.; The food-grade Lactococcus lactis is a potential vector to be used as a live vehicle for the delivery of heterologous proteins for vaccine and pharmaceutical purposes . We constructed a plasmid vector pSVac that harbors a 255-bp single-repeat sequence of the cell wall-binding protein region of the AcmA protein . The recombinant plasmid was transformed into Escherichia coli and expression of the gene fragment was driven by the T7 promoter of the plasmid . SDS-PAGE showed the presence of the putative AcmA' fragment and this was confirmed by Western blot analysis . The protein was isolated and purified using a His-tag affinity column . When mixed with a culture of L . lactis MG1363, ELISA and immunofluorescence assays showed that the cell wall-binding fragment was anchored onto the outer surface of the bacteria . This indicated that the AcmA' repeat unit retained the active site for binding onto the cell wall surface of the L . lactis cells . Stability assays showed that the fusion proteins (AcmA/A1, AcmA/A3) were stably docked onto the surface for at least 5 days . The AcmA' fragment was also shown to be able to strongly bind onto the cell surface of naturally occurring lactococcal strains and Lactobacillus and, with less strength, the cell surface of Bacillus sphericus . The new system designed for cell surface display of recombinant proteins on L . lactis was evaluated for the expression and display of A1 and A3 regions of the VP1 protein of enterovirus 71 (EV71) . The A1 and A3 regions of the VP1 protein of EV71 were cloned upstream to the cell wall-binding domains of AcmA protein and successfully expressed as AcmA/A1 and AcmA/A3 . Whole-cell ELISA showed the successful display of VP1 protein epitopes of EV71 on the surface of L . lactis . The success of the anchoring system developed in this study for docking the A1 and A3 epitopes of VP1 onto the surface of L . lactis cells opens up the possibilities of peptide and protein display for not only Lactococcus but also for other gram-positive bacteria . This novel way of displaying epitopes on the cell surface of L . lactis and other related organisms should be very useful in the delivery of vaccines and other useful proteins. Pol J Vet Sci, 2004, 7(4), 251 - 9 Effect of chosen lactic acid bacteria strains on Staphylococcus aureus in vitro as well as in meat and raw sausages; Gomolka-Pawlicka M et al.; The present study was aimed at determining the influence of 15 strains of lactic acid bacteria on the growth of 2 Staphylococcus aureus strains in vitro as well as in meat and raw sausages . The investigations were performed within the framework of three alternate stages which differed in respect to the products studied, the number of Lactobacillus sp . strains and, partly, methodological approach . The study also considered water activity (a(w)) and pH of the products investigated . The results obtained are demonstrated in 5 diagrams . It was found that among 15 strains of Lactobacillus aureus investigated only one strain, Lactobacillus helveticus T 78, showed antagonistic effect on studied strains of Staphylococcus aureus both in vitro as well as in meat and raw sausages . Five other strains of Lactobacillus spp . displayed the antagonistic effect in vitro only . The temperature and incubation time of sausages, but also the type of sausage stuffing were found to have a distinct or slight influence, respectively, on the antagonistic interaction between the bacteria . However, this phenomenon was affected by neither a(w) nor pH. J Food Prot, 2004 Dec, 67(12), 2779 - 85 Interrelationships among microbiological, physicochemical, and biochemical properties of Terrincho cheese, with emphasis on biogenic amines; Pinho O et al.; Changes in the microbiological, physicochemical, and biochemical characteristics of Terrincho cheese as represented by native microflora, pH, water activity, soluble nitrogen fractions, free amino acids, and biogenic amines (e.g., ethylamine, dimethylamine, tryptamine, phenylethylamine, putrescine, cadaverine, histamine, tyramine, cystamine, and spermine) during ripening were monitored . Terrincho is a traditional Portuguese cheese manufactured from raw ewe's milk . The main groups of microorganisms (lactococci, lactobacilli, enterococci, pseudomonads, staphylococci, coliforms, yeasts, and molds) were determined following conventional microbiological procedures . Free amino acids and biogenic amines were determined by reverse-phase high-performance liquid chromatography, following extraction from the cheese matrix and derivatization with dabsyl chloride . The total content of free amino acids ranged from 1,730 mg/kg of dry matter at the beginning of the ripening stage to 5,180 mg/kg of dry matter by day 60 of ripening; such an increase was highly correlated with the increase of water-soluble nitrogen in total nitrogen, 12% trichloroacetic acid-soluble nitrogen in total nitrogen, and 5% phosphotungstic acid-soluble nitrogen in total nitrogen throughout ripening . Histamine was consistently present at very low levels, whereas putrescine, cadaverine, and tryptamine were the dominant biogenic amines and increased in concentration during ripening . Ethylamine, tryptamine, phenylethylamine, and cystamine reached maxima by 30 days of ripening and decreased thereafter . Significant correlations between amino acid precursors and corresponding biogenic amines, as well as between biogenic amines and microbial viable numbers, were observed. J Food Prot, 2004 Dec, 67(12), 2727 - 34 Characterization and antimicrobial activity of bacteriocin 217 produced by natural isolate Lactobacillus paracasei subsp . paracasei BGBUK2-16; Lozo J et al.; The strain Lactobacillus paracasei subsp . paracasei BGBUK2-16 . which was isolated from traditionally homemade white-pickled cheese, produces bacteriocin 217 (Bac217; approximately 7 kDa) . The onset of Bac217 biosynthesis was observed in the logarithmic phase of growth, and the production plateau was reached after 9 or 12 h of incubation at 37 and 30 degrees C, respectively, when culture entered the early stationary phase . Biochemical characterization showed that Bac217 retained antimicrobial activity within the range of pH 3 to 12 or after treatment at 100 degrees C for 15 min . Bac217 antimicrobial activity also remained unchanged after storage at 4 degrees C for 6 months or -20 degrees C for up to 12 months . However, Bac217 activity was completely lost after treatment with different proteolytic enzymes . BGBUK2-16 contains only one plasmid about 80 kb in size . Plasmid curing indicated that genes coding for Bac217 synthesis and immunity seem to be located on this plasmid . Bac217 exhibited antimicrobial activity against some pathogenic bacteria, such as Staphylococcus aureus and Bacillus cereus . Interestingly, Bac217 showed activity against Salmonella sp . and Pseudomonas aeruginosa ATCC27853 . The inhibitory effect of BGBUK2-16 on the growth of S . aureus in mixed culture was observed . S . aureus treatment with Bac217 led to a considerable decrease (CFU/ml) within a short period of time . The mode of Bac217 action on S . aureus was identified as bactericidal . It should be noted that the strain BGBUK2-16 was shown to be resistant to bacteriocin nisin, which is otherwise widely used as a food additive for fermented dairy products. Home Healthc Nurse, 2005 Jan, 23(1), 13 - 15 Bugs, Drugs, and Yogurt; Neafsey PJ et al.; Mrs . S . is suffering from a yeast infection after a course of antibiotics . Despite several unsuccessful self-treatments with an over-the-counter intravaginal clotrimazole treatment, she remained symptomatic and she was prescribed oral Diflucan (fluconazole) . At first it seems that Mrs . S . has been cured, but a few days later, her symptoms return . You remember reading an article on Lactobacillus acidophilus as a remedy for yeast infections . Is there any validity to this treatment or is it just the latest craze of naturopathic medicine? Appl Microbiol Biotechnol . 2005 Jan 4; {Epub ahead of print} Lactobacillus reuteri ATCC 53608 mdh gene cloning and recombinant mannitol dehydrogenase characterization; Sasaki Y et al.; A gene encoding mannitol-2-dehydrogenase (E.C . 1.1.1.138) (MDH) was cloned from Lactobacillus reuteri and expressed in Escherichia coli . The 1,008-bp gene encodes a protein consisting of 336 amino acids, with a predicted molecular mass of 35,920 Da . The deduced amino acid sequence of L . reuteri MDH (LRMDH) is 77% and 76% similar to the MDHs from Leuconostoc mesenteroides and Leuconostoc pseudomesenteroides, respectively . The purified recombinant enzyme appears as a single band of 40 kDa in sodium dodecyl sulfate-polyacrylamide gel electrophoresis, but gel filtration indicates that the native enzyme is a dimer . The optimum temperature for the recombinant enzyme is 37 degrees C, the pH optima for D: -fructose reduction and D: -mannitol oxidation are 5.4 and 6.2, respectively . The K(m) values for NAD (9 mM) and NADH (0.24 mM) are significantly higher than those for NADP (0.35 mM) and NADPH (0.04 mM) . The K(m) values of LRMDH for D: -fructose and D: -mannitol are 34 mM and 54 mM, respectively . Contrary to what the enzyme sequence suggests, recombinant LRMDH contains a single catalytic zinc per subunit. Biofactors, 2004, 22(1-4), 63 - 6 Antitumor activity and action mechanisms of Lactobacillus casei through the regulation of immune responses; Matsuzaki T et al.; Lactobacillus casei strain Shirota (LcS) has been demonstrated to have beneficial effects in numerous disease models, especially murine autoimmune disease and carcinogenesis models, via host immune modulation . It has also been reported that LcS induced recovery of the host immune responses that were decreased by treatment with carcinogens, and that augmented the natural killer activity and T cell functions of host immune cells . After LcS is ingested by the host, it is incorporated into M cells in Peyer's patches (PP) and digested to form active components . In PP, macrophages or dendritic cells that phagocytosed LcS gained ability to produce tumor necrosis factor-alpha . The components of LcS digested in PP were then recognized through toll-like receptor 2 in macrophages, resulting in the production of several cytokines which elicit varied responses in host immune cells . Also, it was observed by 2D-PAGE analyses that the expression level and/or the phosphorylation of some proteins in PP and mesenteric lymph nodes were definitely altered after the ingestion of LcS, providing more evidence of cellular responses. Biofactors, 2004, 21(1-4), 155 - 8 Isolation and identification of lactic acid bacteria with effect of immune protection to Eschericia coli in mice; Ishida-Fujii K et al.; Lactic acid bacteria were isolated from an alcohol fermentation broth, and the activity as a probiotic was examined using pathogenic E . coli . Thirty-six strains exhibiting good growth were isolated in the medium of concentrated mush which was a residue resulted in the alcohol distillation process . One of these strains, Lactobacillus paracasei subsp . paracasei I-5, could be grown in the medium containing 8 vol% ethanol and at 45 degrees C . The characteristics were different from the type strain, L . paracasei subsp . paracasei NBRC 15889trade mark . L . paracasei I-5 showed an excellent growth in the concentrated mush, which just diluted two-fold and adjusted the pH . ICR mice were fed with a standard germ-free feed (CMF) and the strain I-5 (7x10(9) cells/day) was orally administrated for 11 days prior to the intraperitoneal challenge with pathogenic E . coli Juhl . After the challenge, mice administrated the strain I-5 exhibited a high survival rate and survival extension days (p < 0.01) compared with the control . The results suggested that the strain might enhance the animal resistance against microbial pathogens . Neonatal diarrhea caused by E . coli is a serious disease in calf breeding . The strain might be practically valuable to prevent diarrhea in calves. Pediatrics, 2005 Jan, 115(1), 178 - 81 Lactobacillus sepsis associated with probiotic therapy; Land MH et al.; Probiotic strains of lactobacilli are increasingly being used in clinical practice because of their many health benefits . Infections associated with probiotic strains of lactobacilli are extremely rare . We describe 2 patients who received probiotic lactobacilli and subsequently developed bacteremia and sepsis attributable to Lactobacillus species . Molecular DNA fingerprinting analysis showed that the Lactobacillus strain isolated from blood samples was indistinguishable from the probiotic strain ingested by the patients . This report indicates, for the first time, that invasive disease can be associated with probiotic lactobacilli . This report should not discourage the appropriate use of Lactobacillus or other probiotic agents but should serve as a reminder that these agents can cause invasive disease in certain populations. Vaccine, 2005 Jan 19, 23(9), 1126 - 34 Intranasal immunisation using recombinant Lactobacillus johnsonii as a new strategy to prevent allergic disease; Scheppler L et al.; We have previously demonstrated the induction of a specific anti-IgE response in vivo by parenteral immunisation of rhesus monkeys using short IgE mimotopes or an anti-idiotypic antibody mimicking an IgE epitope . Such specific anti-IgE responses may be of clinical benefit for atopic patients . In this study, we examined the potential for a more convenient therapy via mucosal immunisation using live recombinant Lactobacillus johnsonii (Lj) as a vaccine delivery vehicle . Either an anti-idiotypic scFv or an IgE mimotope were expressed on the surface of Lj as fusion proteins using the cell wall anchored proteinase PrtB from Lactobacillus delbrueckii subsp . bulgaricus . The recombinant Lj were shown to express the heterologous fusion proteins and were specifically recognised by the corresponding anti-human IgE monoclonal antibody . Subcutaneous and intranasal immunisation of mice with recombinant Lj, expressing these fusion proteins induced a systemic IgG response against human IgE . Our data suggest that recombinant Lactobacilli expressing IgE epitopes may represent a novel means of vaccination to induce a beneficial anti-IgE response. Biopolymers . 2004 Dec 29;77(2):98-106 {Epub ahead of print} Viscometric and static light scattering studies on an exopolysaccharide produced by Lactobacillus delbrueckii subspecies bulgaricus NCFB 2483; Goh KK et al.; The rheological properties and molecular parameters of a purified exopolysaccharide (EPS) produced by a ropy strain of Lactobacillus delbrueckii subsp . bulgaricus NCFB 2483 were investigated . Using capillary viscometry, an intrinsic viscosity of 2,013 mL/g was obtained . The flow curves were fitted by both the Carreau and the Cross equations for shear-thinning fluids, with the Carreau equation giving a better fit . The Cross equation fitted fairly well the plot of reduced viscosity as a function of reduced shear rate with an exponent value (1 - n) of approximately 0.76, typical of random coil polymers . Furthermore, the concentration dependence of the viscosity plot showed a gradient of approximately 1.1 in the dilute regime and 3.3 in the semidilute regime . Molecular parameters were obtained using a multiangle laser light scattering technique . The 2483 EPS molecules had a weight-average molar mass of approximately 2 x 10(6) Da and a z-average root mean square radius (RMS) of approximately 151 nm . From the light scattering data, the bacterial EPS was also found to have a low polydispersity index ( approximately 1.15) and adopt a random coil conformation . (c) 2004 Wiley Periodicals, Inc . Biopolymers, 2005. Nutr Clin Care, 2004 Jul-Sep, 7(3), 104 - 13 Probiotics and prebiotics in clinical practice; Broussard EK et al.; Probiotics are living organisms that have a beneficial therapeutic effect on the host when ingested . The bacteria Lactobacillus GG and the yeast Saccharomyces boulardii have been shown in controlled trials to prevent antibiotic-associated diarrhea . Prebiotics are non-living, indigestible polysaccharides metabolized in the intestine that induce physical and bacteriologic musocal modifications . Rather than introducing exogenous bacteria into the intestinal millieu, prebiotics stimulate certain indigenous bacteria already resident in the gut. Biosci Biotechnol Biochem, 2004 Dec, 68(12), 2427 - 35 Isolation of Lactobacillus sakei Strain KJ-2008 and Its Removal of Characteristic Malodorous Gases under Anaerobic Culture Conditions; Kim JD et al.; A number of different sources, such as composts, leachates, and pig feces samples were collected from different pig farms in Korea . Several microorganisms were screened for their ability to deodorize the malodorous gases . As a result, a novel malodorous gas-deodorizing bacterial strain KJ-2008 was isolated due to the most abundant of nitrate-supplemented minimal media under anaerobic conditions . Crimp-sealed serum bottles containing nitrate-supplemented minimal medium (MM-NO(3)(-)) in airtight conditions were inoculated with KJ-2008 . Nitrate concentration decreased rapidly after 20 h incubation and nitrite production reached almost zero during the time the experimental was carried out . Taxonomic identification including 16S rDNA base sequencing and phylogenetic analysis indicated that the isolate KJ-2008 had a 99.8% homology in its 16S rDNA base sequence with Lactobacillus sakei . Among the volatile fatty acids, acetic acid contained in large amounts in fresh piggery slurry decreased about 40% after 50 h incubation of the strain KJ-2008 . n-Butyric acid, n-valeric acid, and iso-valeric acid gradually decreased, and iso-butyric acid and capronic acid dramatically eliminated at initial time with the treatment . Moreover, NH(3) removal efficiency reached a maximum of 98.5% after 50 h of incubation . The concentration of H(2)S did not change. Infect Immun, 2005 Jan, 73(1), 476 - 84 CCL20/macrophage inflammatory protein 3alpha and tumor necrosis factor alpha production by primary uterine epithelial cells in response to treatment with lipopolysaccharide or Pam3Cys; Crane-Godreau MA et al.; Having previously shown that CCL20/macrophage inflammatory protein 3alpha and tumor necrosis factor alpha (TNF-alpha) are released by polarized primary rat uterine epithelial cells (UEC) in response to Escherichia coli but not to Lactobacillus rhamnosus, we sought to determine if epithelial cells are responsive to pathogen-associated molecular patterns (PAMP), including lipopolysaccharide (LPS), lipoteichoic acid (LTA), and Pam(3)Cys, a bacterial lipoprotein analog . Epithelial cells were grown to confluence on Nunc cell culture inserts prior to apical treatment with PAMPs . In response to LPS, LTA, and Pam(3)Cys (EMC Microcollection GmbH, Tubingen, Germany), CCL20 levels increased (4- to 10-fold) while PAMPs caused increased TNF-alpha (1- to 4-fold) in the medium collected after 24 h of incubation . Both apical and basolateral secretion of CCL20 and TNF-alpha increased in response to PAMPs, but treatments had no effect on cell viability and integrity, as measured by transepithelial resistance . Time course studies of CCL20 and TNF-alpha release in response to Pam(3)Cys and LPS indicated that CCL20 release peaked between 2 and 4 h after treatment, whereas TNF-alpha release was gradual over the length of the incubation . Freeze-thaw and cell lysis experiments, along with actinomycin D studies, suggested that CCL20 and TNF-alpha are synthesized in response to PAMP stimulation . Taken together, these studies demonstrate that E . coli and selected PAMPs have direct effects on the production of CCL20 and TNF-alpha without affecting cell integrity . Since CCL20 is known to be both chemotactic and antimicrobial, the increase in apical and basolateral release by UEC in response to PAMPs suggests a new mechanism of innate immune protection in the female reproductive tract. Int J Food Microbiol, 2005 Jan 15, 98(1), 81 - 8 Influence of polysaccharides on oxygen dependent lactate utilization by an amylolytic Lactobacillus plantarum strain; Pintado J et al.; Oxygen-dependent conversion of lactate to acetate by the amylolytic strain Lactobacillus plantarum A6 was studied using MRS-grown cells, transferred to a basic medium with lactate . In the presence of oxygen, lactate was stoechiometricaly converted to acetate . When glucose, maltose or cellobiose was added to the basic medium, no utilisation of lactate was observed . However, when starch or glycogen was added, the conversion of lactate to acetate happened . To verify the possible link of this effect with sugar consumption rate, a glucose-fed batch culture was conducted with a lactic acid consuming culture grown on the basic medium with lactate . Even when glucose was fed at the same low rate as the consumption rate observed for polysaccharides, lactic acid was no more consumed . For the amylolytic strain L . plantarum A6, the transport and use of oligosaccharides resulting from polysaccharides hydrolysis might affect differently the glycolytic flux, with the putative consequence to suppress the metabolic control of glycolysis by glucose . This fact could play an important role in the fermentation of amylaceous foods. Int J Food Microbiol, 2005 Jan 15, 98(1), 63 - 72 Phenotypic characterization of lactic acid bacteria from sourdoughs for Altamura bread produced in Apulia (Southern Italy); Ricciardi A et al.; In order to study the composition of the lactic acid bacteria (LAB) community of sourdoughs used for the manufacture of Altamura bread, a traditional durum wheat bread produced in Apulia (Southern Italy), 111 strains of LAB were isolated and characterized . The phenotypic characterization of the isolates, carried out using a set of 29 tests, allowed the identification of 15 clusters at the 80% similarity level by hierarchical cluster analysis . Of the isolates, 88% were identified as facultatively heterofermentative LAB (Lactobacillus plantarum, Lb . paracasei, Lb . casei) and 12% as heterofermentative LAB (Lb . brevis, Leuconostoc mesenteroides) . SDS-PAGE profiles of whole cell proteins of 68 strains confirmed the identification . Both the diversity and structure of the lactic microflora for sourdoughs for Altamura bread varied among samples. Int J Food Microbiol, 2005 Jan 15, 98(1), 53 - 62 A putative glucan synthase gene dps detected in exopolysaccharide-producing Pediococcus damnosus and Oenococcus oeni strains isolated from wine and cider; Walling E et al.; Some lactic acid bacteria can induce viscosity in wine, beer and cider by production of exopolysaccharides (EPS) . A polymerase chain reaction (PCR) assay was previously described for the detection of ropy Pediococcus damnosus strains in wine {J . Appl . Microbiol . 90 (2001) 535} . The primers used in that study, PF5 and PF6, are investigated in addition to new primers which broaden the range of spoiling agents detectable by PCR . Primers PF1 and PF8 allow the amplification of DNA from ropy P . damnosus strains isolated from wine, as was observed with PF5 and PF6 . In addition, PF1 and PF8, unlike PF5 and PF6, are able to generate an amplicon using template DNA from a ropy P . damnosus strain isolated from cider and a ropy Oenococcus oeni strain isolated from champagne . Two different ropy Lactobacillus species were also isolated, but their DNA was not amplified using primers PF1 and PF8 . The new primers PF1 and PF8 were chosen from the sequence of gene dps, a putative glucan synthase gene, found across all the ropy P . damnosus strains isolated, from both wine or cider, and also in a ropy O . oeni strain . To our knowledge, this is the first time that an EPS-producing O . oeni strain is described . Glucan biosynthesis was assessed by agglutination tests done with Streptococcus pneumoniae type 37-specific antibodies, which specifically detect glucan-producing cells . The results show that there is a direct correlation between glucan production and detection of gene dps . Therefore, Dps is considered a candidate for the glucan synthase enzyme responsible for EPS production by ropy strains of P . damnosus and O . oeni. Antimicrob Agents Chemother, 2005 Jan, 49(1), 170 - 5 Ecological effects of perorally administered pivmecillinam on the normal vaginal microflora; Sullivan A et al.; The knowledge of the effects of antimicrobial agents on the normal vaginal microflora is limited . The objective of the present study was to study the ecological impact of pivmecillinam on the normal vaginal microflora . In 20 healthy women, the estimated day of ovulation was determined during three subsequent menstrual cycles . Microbiological and clinical examinations were performed on the estimated day of ovulation and on day 3 in all cycles and also on day 7 after ovulation in cycles 1 and 2 . Anaerobic and facultative anaerobic gram-positive rods, mainly species of lactobacilli and actinomycetes, dominated the microflora . One woman was colonized on the third day of administration with a resistant Escherichia coli strain, and Candida albicans was detected in one woman on days 3 and 7 in cycle 2 . No other major changes in the normal microflora occurred during the study . Administration of pivmecillinam had a minor ecological impact on the normal vaginal microflora. Genet Mol Res, 2004 Sep 30, 3(3), 432 - 40 Reduction of non-digestible oligosaccharides in soymilk: application of engineered lactic acid bacteria that produce alpha-galactosidase; LeBlanc JG et al.; Human consumption of soy-derived products has been limited by the presence of non-digestible oligosaccharides (NDO), such as the alpha-galactooligosaccharides raffinose and stachyose . Most mammals, including man, lack pancreatic alpha-galactosidase (alpha-Gal), which is necessary for the hydrolysis of these sugars . However, such NDO can be fermented by gas-producing microorganisms present in the cecum and large intestine, which in turn can induce flatulence and other gastrointestinal disorders in sensitive individuals.The use of microorganisms expressing alpha-Gal is a promising solution to the elimination of NDO before they reach the large intestine . In the present study, lactic acid bacteria engineered to degrade NDO have been constructed and are being used as a tool to evaluate this solution . The alpha-Gal structural genes from Lactobacillus plantarum ATCC8014 (previously characterized in our laboratory) and from guar have been cloned and expressed in Lactococcus lactis . The gene products were directed to different bacterial compartments to optimize their possible applications . The alpha-Gal-producing strains are being evaluated for their efficiency in degrading raffinose and stachyose: i) in soymilk fermentation when used as starters and ii) in situ in the upper gastrointestinal tract when administered to animals orally, as probiotic preparations . The expected outcomes and possible complications of this project are discussed. Appl Microbiol Biotechnol . 2004 Dec 22; {Epub ahead of print} Production of Lactobacillus kefir cells for asymmetric synthesis of a 3,5-dihydroxycarboxylate; Pfruender H et al.; An efficient fedbatch process for the production of Lactobacillus kefir DSM 20587 cells was developed . An improvement in space time yield of 270% (3.7 g(DCW) l(-1) day(-1)) and in final enzyme activity of 440% (9.1 U/ml) was achieved on a 150 l scale by controlling the oxygen transfer rate to 7-9 mmol l(-1) h(-1) . The cells exhibited good and highly stereoselective reducing activities against tert-butyl 6-chloro-3,5-dioxohexanoate . tert-Butyl (3R,5S)-6-chloro-dihydroxyhexanoate, a chiral building block for HMG-CoA reductase inhibitor synthesis, was produced with 47.5% yield and >99% ee at C(3)3 and C(5)5 in a simple batch biotransformation process. Lett Appl Microbiol, 2005, 40(1), 56 - 62 Gene organization and sequences of pediocin AcH/PA-1 production operons in Pediococcus and Lactobacillus plasmids; Miller KW et al.; Abstract k.w . miller, p . ray, t . steinmetz, t . hanekamp and b . ray 2004.Aims: To determine the locations and sequences of pediocin AcH production genes in Pediococcus parvulus ATO77 from vegetables, Lactobacillus plantarum WHE92 from Muenster cheese, and a lactose-fermenting isolate Pediococcus pentosaceus S34 from buffalo milk . Methods and Results: Plasmid curing, Southern blot hybridization, and DNA sequence analysis indicate that pediocin AcH production genes are encoded by highly similar operons in unique plasmids designated pATO77 from P . parvulus ATO77, pS34 from P . pentosaceus S34, and pWHE92 from Lact . plantarum WHE92 . Structure, immunity and secretion system genes are linked together in the operons, and the promoter sequences are the same . The amino acid sequences of the encoded proteins are highly conserved between plasmids . Conclusions: Pediocin AcH production genes are located within a plasmid-borne operon cassette in all lactic acid bacterial strains examined to date . All four genes needed for production are present within a single plasmid in each strain . Significance and Impact of the Study: This is the first demonstration that the expression of a class IIa bacteriocin is directed by a common gene cassette that has been disseminated to unique plasmids in different genera of lactic acid bacteria . These plasmids should be useful for expressing pediocin AcH in Pediococcus and Lactobacillus strains used in food production. J Agric Food Chem, 2004 Dec 29, 52(26), 7752 - 8 High-performance liquid chromatographic determination of naturally occurring folates during tempe preparation; Ginting E et al.; A trienzyme treatment (protease, alpha-amylase, and human plasma conjugase), followed by purification using SPE with SAX cartridges and reversed-phase HPLC with UV-PDA detection, was performed for determination of the distribution of various folate forms and content at various stages of tempe preparation . The major folate form in soybean identified was 5-formyl tetrahydrofolate (5-CHO-H4folate), followed by 10-formyl tetrahydrofolate (10-CHO-PGA), and 5-methyl tetrahydrofolate (5-CH3-H4folate), whereas folic acid was not detected and tetrahydrofolic acid (H4folate) was not detectable . The most predominant form in tempe was also 5-CHO-H4folate, followed by 10-CHO-PGA, whereas the quantities of 5-CH3-H4folate and folic acid were negligible . Quantities and retention of folate significantly decreased during the first boiling, dehulling, soaking, and second boiling procedures, yielding folate retention of 32% . A remarkable increase in folate content was found after fermentation, 5.2-fold higher than that of the boiled soybean . This may be due to de novo formation of folate by Rhizopus oligosporus, the principal mold in tempe fermentation . HPLC results were approximately 38-55% lower than the values obtained from the microbiological assay using Lactobacillus casei. J Appl Microbiol, 2005, 98(1), 229 - 37 Characterization of an acid phosphatase from Lactobacillus pentosus: regulation and biochemical properties; Palacios MC et al.; Abstract m.c . palacios, m . haros, c.m . rosell and y . sanz . 2004.Aims: To screen for phosphatase and phytase activities in Lactobacillus isolated from diverse ecosystems and to determine the biochemical properties and the factors that regulate the synthesis of the enzyme responsible for these activities in the selected strain, Lactobacillus pentosus CECT 4023 . Methods and Results: These activities were determined spectrophotometrically by using p-nitrophenyl phosphate and sodium phytate as substrates . They were maximal at the onset of the stationary phase of growth and repressed in the presence of high glucose concentration and inorganic phosphate . The enzyme responsible for these activities was an acid phosphatase (E.C.3.1.3.2.), with a molecular mass of 69 kDa . The activity was optimum at pH 5.0 and 50 degrees C . It hydrolysed mono-phosphorylated substrates and phytate, albeit at lower rates . It was inhibited by iodoacetic acid, phenyl-methylsulphonyl fluoride, di-sodium pyrophosphate and Ca(+2) while activated by Co(+2) and low concentrations of l-ascorbic acid and EDTA . Conclusions: Lactobacillus pentosus CECT 4023 produces a nonspecific acid phosphatase that hydrolyses a number of mono-phosphorylated substrates and phytate . Significance and Impact of the Study: The results suggest that the phosphatase from L . pentosus CECT 4023 could partly contribute to reduce the phosphorylation degree of phytate and its derivatives and, thereby, their anti-nutrient properties during fermentation processes. J Appl Microbiol, 2005, 98(1), 84 - 95 Cell wall modifications during osmotic stress in Lactobacillus casei; Piuri M et al.; Abstract m . piuri, c . sanchez-rivas and s.m . ruzal . 2004.Aims: To study the modification of the cell wall of Lactobacillus casei ATCC 393 grown in high salt conditions . Methods and Results: Differences in the overall structure of cell wall between growth in high salt (MRS + 1 mol l(-1) NaCl; N condition) and control (MRS; C condition) conditions were determined by transmission electronic microscopy and analytical procedures . Lactobacillus casei cells grown in N condition were significantly larger than cells grown under unstressed C condition . Increased sensitivity to mutanolysin and antibiotics with target in the cell wall was observed in N condition . Purified cell wall also showed the increased sensitivity to lysis by mutanolysin . Analysis of peptidoglycan (PG) from stressed cells showed that modification was at the structural level in accordance with a decreased PG cross-link involving penicillin-binding proteins (PBP) . Nine PBP were first described in this species and these proteins were expressed in low percentages or presented a modified pattern of saturation with penicillin G (Pen G) during growth in high salt . Three of the essential PBP were fully saturated in N condition at lower Pen G concentrations than in C condition, suggesting differences in functionality in vivo . Conclusions: The results show that growth in high salt modified the structural properties of the cell wall . Significance and Impact of Study: Advances in understanding the adaptation to high osmolarity, in particular those involving sensitivity to lysis of lactic acid bacteria. J Appl Microbiol, 2005, 98(1), 64 - 72 Analysis of promoter sequences from Lactobacillus helveticus CNRZ32 and their activity in other lactic acid bacteria; Chen YS et al.; Abstract y.-s . chen and j.l . steele . 2004.Aims: To clone and analyse seven putative promoter fragments (pepC, pepN, pepX, pepO, pepE, pepO2, hsp17) from Lactobacillus helveticus CNRZ32 for their expression in Lact . helveticus CNRZ32, Lact . casei ATCC334 and Lactococcus lactis MG1363 . Methods and Results: Promoter fragments were fused to the promoter-less beta-glucuronidase (gusA) gene on pNZ272(RBS-) (ATG-) . The resulting constructs were evaluated for their ability to drive the expression of active GusA with 0.5 mmol l(-1) 5-bromo-4-chloro-3-indolyl-beta-d-glucuronide . All promoters except P(pepN)::gusA were active in the examined strains . Northern hybridization was performed to examine the promoter strength . Sequence analysis of these promoters identified well conserved putative ribosomal binding and putative -10 hexamers sites . Conclusions: Seven promoter fragments from Lact . helveticus CNRZ32 were recognized in the lactic acid bacteria, Lact . casei ATCC334 and L . lactis MG1363, as well as in Escherichia coli . P(pepN)::gusA could not be maintained in the strains examined because of toxicity associated with heterologous protein over-expression driven by P(pepN) . Significance and Impact of the Study: This study revealed that desirable levels of heterologous food-grade protein production in GRAS organisms can be obtained with the application of natural promoter fragments from closely related organisms. J Appl Microbiol, 2005, 98(1), 56 - 63 Lyophilized preparations of bacteriocinogenic Lactobacillus curvatus and Lactococcus lactis subsp . lactis as potential protective adjuncts to control Listeria monocytogenes in dry-fermented sausages; Benkerroum N et al.; Abstract n . benkerroum, a . Daoudi, t . hamraoui, h . Ghalfi, c . thiry, m . duroy, p . evrart, d . roblain and p . thonart . 2004.Aim: Study of the effectiveness of in situ bacteriocin production by lactic acid bacteria (LAB) to control Listeria monocytogenes in dry-fermented sausages . Methods and Results: Two bacteriocin-producing strains: Lactococcus lactis subsp . lactis LMG21206 and Lactobacillus curvatus LBPE were grown in a pilot scale fermentor and lyophilized to be directly used in dry sausage fermentation . A commercial starter culture (Bel'meat(TM) SL-25) not inhibitory to L . monocytogenes (Bac(-) starter) was mixed (1 : 1) with each of the two lyophilized bacteriocin-producing strains to obtain starters active against the pathogen (Bac(+) starter) . Anti-Listeria effectiveness of the Bac(+) starters was studied in dry-fermented sausages . The meat batter was experimentally contaminated with a mixture of four different strains of L . monocytogenes (10(2)-10(3) CFU g(-1)) . The results showed that L . monocytogenes did not grow in any of the contaminated batches, but no significant decrease (P > 0.05) was observed either in the positive control (no added starter culture) or in samples fermented with the Bac(-) starter culture during the fermentation period and up to 15 days of drying . When the Bac(+) starter contained Lb . curvatus LBPE, cell counts of L . monocytogenes decreased to below the detectable limit (<10 CFU g(-1)) after 4 h of fermentation and no survivors could be recovered by enrichment beyond day 8 of drying . When the Bac(+) starter culture containing Lc . lactis LMG21206 was used, a decrease in Listeria counts to below the detectable limit was achieved after 15 days of drying . Conclusions: The bacteriocin-producing strains studied may be used as adjunct cultures for sausage fermentations to control the occurrence and survival of L . monocytogenes . Significance and Impact of the Study: Addition of the Bac(+) strains, especially the Lb . curvatus strain would provide an additional hurdle to enhance the control of L . monocytogenes in fermented meat products. J Appl Microbiol, 2005, 98(1), 14 - 23 Succinate production and citrate catabolism by Cheddar cheese nonstarter lactobacilli; Dudley EG et al.; Abstract e.g . dudley and j.l . steele . 2004.Aims: To identify strains of Cheddar cheese nonstarter lactobacilli that synthesize succinate from common precursors and characterize the biochemical pathways utilized . Methods and Results: Whole cell incubations of Lactobacillus plantarum, Lactobacillus casei, Lactobacillus zeae and Lactobacillus rhamnosus, were used to identify strains that accumulated succinate from citrate, l-lactate, aspartic acid or isocitrate . In vivo(13)C-nuclear magnetic resonance spectroscopy ((13)C-NMR) identified the biochemical pathway involved at pH 7.0, and under conditions more representative of the cheese ripening environment (pH 5.1/4% NaCl/13 degrees C) . Enzyme assays on cell-free extracts were used to support the pathway suggested by (13)C-NMR . Conclusions: The Lact . plantarum strains studied synthesize succinate from citrate by the reductive tricarboxylic acid (TCA) cycle at either pH 7.0 or pH 5.1/4% NaCl/13 degrees C . Lactobacillus casei, Lact . zeae and Lact . rhamnosus strains lack one or more enzymatic activities present in this pathway, and do not accumulate succinate from any of the four precursors studied . Significance and Impact of the Study: The addition of Lact . plantarum strains to milk during cheese manufacture may increase the accumulation of the flavour enhancer succinate. Ann Allergy Asthma Immunol, 2004 Dec, 93(6), 601 - 5 Rice-induced enterocolitis in an infant: TH1/TH2 cellular hypersensitivity and absent IgE reactivity; Gray HC et al.; BACKGROUND: Although food allergy is common in children, rice allergy is unusual in Western cultures . OBJECTIVE: To report a case of T-cell-mediated rice intolerance in an 11-month-old girl . METHODS: To evaluate the intolerance to rice in this patient, a graded rice food challenge was performed . To examine the immunologic reactivity to rice, in vitro lymphoproliferative responses and cytokine synthesis of rice-stimulated peripheral blood lymphocytes (PBLs) was performed . Subsequently, skin patch testing to rice and other foods was performed . RESULTS: Allergy skin prick test results were negative for rice and positive for egg, milk, and soy . Specific IgE antibodies to rice, egg, peanut, wheat, walnut, codfish, milk, soybean, corn, shrimp, scallops, and clams were undetectable . Results of a single-blind rice food challenge were positive, manifested by emesis that persisted for more than an hour and required intravenous hydration . In vitro lymphoproliferation by the patient's PBLs to rice stimulation was positive . In addition, cytokine synthesis of interferon-gamma, interleukin 10 (IL-10), tumor necrosis factor a, and IL-5 by the patient's rice-stimulated PBLs was elevated, indicating a TH1/TH2 cell response to rice . Endoscopy revealed normal esophageal, gastric, and duodenal mucosa; a biopsy specimen revealed mild esophagitis . Duodenal explant T cells were initially established by stimulation with rice and IL-2 . After a 2-day rest, the lymphocytes were restimulated with rice for 7 days and revealed increased interferon-gamma and IL-5 synthesis . Twenty billion colony forming units of Lactobacillus GG were added to the patient's diet twice daily . After 6 weeks, rice rechallenge resulted in emesis within 1 hour . Results of patch testing were positive to rice, wheat, and barley but negative to soy, which the patient tolerated on food challenge . CONCLUSIONS: Although this patient did not demonstrate IgE antibody to rice, TH1/TH2 cell-mediated responses to rice were detected, and the patient experienced significant morbidity . Patch testing for gastrointestinal food allergies may be useful when the food specific IgE antibody is negative . Probiotic therapy in this patient did not ameliorate her sensitivity to rice, and food elimination remains the only reliable treatment for TH1/TH2-mediated food hypersensitivity. J Dairy Res, 2004 Nov, 71(4), 461 - 70 Amino acid transamination in permeabilised cells of Lactobacillus helveticus, Lb . paracasei and Lb . danicus; Thage BV et al.; Aminotransferase (AT) activity against 18 amino acids was studied in ten strains of three species of Lactobacillus . A method for permeabilisation of cells was developed using toluene and ethanol combined with mechanical treatment . It was found that the AT activities in the washed permeabilised cells (W-PC) corresponded well to that in cell-free extracts (CFE) . The AT specificity pattern was species as well as strain dependant . Strains of Lb . helveticus had high specificity for aromatic amino acids (ArAA) and lower activity against branched-chain amino acids (BcAA) and Asp, while strains of Lb . paracasei subsp . paracasei degraded BcAA and Asp, but had a lower and variable specificity against ArAA . One of the Lb . paracasei strains was characterised by having very high AT activity against all three BcAA (Ile, Leu, Val) compared with any of the other Lb . paracasei strains tested . Strains of Lb . danicus, which is a newly discovered Lactobacillus species isolated from cheese, had up to about 20 times higher AT activity against Leu than Lb . paracasei and Lb . helveticus . The permeabilised cells of Lb . danicus had also considerably higher AT activity against ArAA than Lb . paracasei and Lb . helveticus strains, and also higher AT activity against Asp . All Lactobacillus strains tested had AT activity against Met, but at a much lower rate than against other amino acids . Results of this study also demonstrated a chemical reaction between alpha-ketoglutaric acid and Asp that was catalysed by pyridoxal-5-phosphate without any AT present. J Dairy Res, 2004 Nov, 71(4), 451 - 60 In vitro screening of potential probiotic activities of selected lactobacilli isolated from unpasteurized milk products for incorporation into soft cheese; Coeuret V et al.; The aim was to select potentially probiotic lactobacilli from 88 strains isolated from unpasteurized milk and cheese products, and to incorporate these bacteria in a viable state into a soft cheese, without changing its quality . The survival of these bacteria was assessed in acidic and bile conditions, after freezing at -80 degrees C . Four strains from unpasteurized Camembert--two Lactobacillus plantarum strains and two Lb . paracasei/casei strains--were identified and typed by PCR and PFGE and were found to display potentially probiotic characteristics in addition to resistance to low pH and bile . These characteristics were resistance to lysozyme, adhesion to CACO-2 cells, antimicrobial effects against common foodborne pathogens (Listeria monocytogenes, Staphylococcus aureus, Salmonella spp., Escherichia coli, innocuity following the ingestion of high doses by mice and appropriate antibiotic susceptibility profiles . The potential of Lb . plantarum strain UCMA 3037 for incorporation into a soft cheese (Pont-l'Eveque registered designation of origin (RDO)) was investigated . This strain grew well and survived in sufficient numbers (more than 10(7) cfu/g throughout the shelf-life of the product) in the cheese . This strain did not change the quality score of the product until the best before date (75 days after manufacture) . Thus, unpasteurized Camembert is a natural source of potentially probiotic lactobacilli, which could be used as an additive in the development of potentially probiotic soft cheeses . Further work is required to demonstrate the persistence and efficacy of these strains in the human host upon ingestion. Biotechnol Lett, 2004 Nov, 26(22), 1739 - 44 Improved synthesis of chiral alcohols with Escherichia coli cells co-expressing pyridine nucleotide transhydrogenase, NADP(+)-dependent alcohol dehydrogenase and NAD(+)-dependent formate dehydrogenase; Weckbecker A et al.; Recombinant pyridine nucleotide transhydrogenase (PNT) from Escherichia coli has been used to regenerate NAD(+) and NADPH . The pnta and pntb genes encoding for the alpha- and beta-subunits were cloned and co-expressed with NADP(+)-dependent alcohol dehydrogenase (ADH) from Lactobacillus kefir and NAD(+)-dependent formate dehydrogenase (FDH) from Candida boidinii . Using this whole-cell biocatalyst, efficient conversion of prochiral ketones to chiral alcohols was achieved: 66% acetophenone was reduced to ( R )-phenylethanol over 12 h, whereas only 19% ( R )-phenylethanol was formed under the same conditions with cells containing ADH and FDH genes but without PNT genes . Cells that were permeabilized with toluene showed ketone reduction only if both cofactors were present. Biotechnol Lett, 2004 Oct, 26(20), 1613 - 6 Fermentative production of DL-lactic acid from amylase-treated rice and wheat brans hydrolyzate by a novel lactic acid bacterium, Lactobacillus sp; Yun JS et al.; Rice and wheat brans, without additional nutrients and hydrolyzed by alpha-amylase and amyloglucosidase, were fermented to DL-lactic acid using a newly isolated strain of Lactobacillus sp . RKY2 . In batch fermentations at 36 degrees C and pH 6, the amount of lactic acid in fermentation broth reached 129 g l(-1) by supplementation of rice bran with whole rice flour . The maximum productivity was 3.1 g lactic acid l(-1) h(-1) in rice bran medium supplemented with whole rice flour or whole wheat flour. Ann N Y Acad Sci, 2004 Nov, 1026, 284 - 8 Immunization of Bovines with Concealed Antigens from Haematobia irritans; Bautista CR et al.; To evaluate an immunization procedure using antigens from Haematobia irritans intestine (AgHiI), four bovines (group I) were inoculated with AgHiI mixed with Freund's incomplete adjuvant containing Lactobacillus casei, three bovines (group II) received AgHiI, and three bovines (group III) received saline solution . At day 35, blood was collected from each animal to feed H . irritans flies . There was no difference in the fly mortality observed in the three groups . The percentage of reduction of eggs oviposited by each female in 8 days (%RE), as compared with group III, was 29.45 for group I and 11.02 for group II . Antibody levels (AbL) to AgHiI were higher in group I than in groups II and III . A high correlation between %RE and AbL was observed. Stomatologiia (Mosk), 2004, 83(6), 6 - 8 {Development and use of periodontal dressing of collagen and Lactobacillus casei 37 cell suspension in combined treatment of periodontal disease of inflammatory origin (results of microbiologic study)}; Effect of yeast inoculation rate on the metabolism of contaminating lactobacilli during fermentation of corn mash; Alltech Biotechnology Center, 3031 Catnip Hill Pike, Nicholasville, KY, 40356, USA, nnarendranath@alltech.comTwo separate 4 (bacterial concentrations)x6 (yeast concentrations) full factorial experiments were conducted in an attempt to identify a novel approach to minimize the effects caused by bacterial contamination during industrial production of ethanol from corn . Lactobacillus plantarum and Lactobacillus paracasei, commonly occurring bacterial contaminants in ethanol plants, were used in separate fermentation experiments conducted in duplicate using an industrial strain of Saccharomyces cerevisiae, Allyeast Superstart . Bacterial concentrations were 0, 1x10(6), 1x10(7) and 1x10(8) cells/ml mash . Yeast concentrations were 0, 1x10(6), 1x10(7), 2x10(7), 3x10(7), and 4x10(7) cells/ml mash . An increased yeast inoculation rate of 3x10(7) cells/ml resulted in a greater than 80% decrease (P<0.001) and a greater than 55% decrease (P<0.001) in lactic acid production by L . plantarum and L . paracasei, respectively, when mash was infected with 1x10(8) lactobacilli/ml . No differences (P>0.25) were observed in the final ethanol concentration produced by yeast at any of the inoculation rates studied, in the absence of lactobacilli . However, when the mash was infected with 1x10(7) or 1x10(8) lactobacilli/ml, a reduction of 0.7-0.9% v/v (P<0.005) and a reduction of 0.4-0.6% v/v (P<0.005) in the final ethanol produced was observed in mashes inoculated with 1x10(6) and 1x10(7) yeast cells/ml, respectively . At higher yeast inoculation rates of 3x10(7) or 4x10(7) cells/ml, no differences (P>0.35) were observed in the final ethanol produced even when the mash was infected with 1x10(8) lactobacilli/ml . The increase in ethanol corresponded to the reduction in lactic acid production by lactobacilli . This suggests that using an inoculation rate of 3x10(7) yeast cells/ml reduces the growth and metabolism of contaminating lactic bacteria significantly, which results in reduced lactic acid production and a concomitant increase in ethanol production by yeast. Eur J Clin Microbiol Infect Dis . 2004 Dec 14; {Epub ahead of print} Pathogenic relevance of Lactobacillus: a retrospective review of over 200 cases; Cannon JP et al.; Given that Lactobacillus has been reported to be the causative pathogen in many types of infection despite debate regarding the organism's clinical significance, a literature review was conducted to investigate the treatments and outcomes of Lactobacillus infections reported to date . In this article, the characteristics of over 200 reported cases of Lactobacillus-associated infections are summarized . Lactobacillus was found to be frequently associated with endocarditis and bacteremia . Lactobacillus was also associated with a variety of other infections including, but not limited to, peritonitis, abscesses, and meningitis . The species casei and rhamnosus were the most common . The isolates tended to be most sensitive to erythromycin and clindamycin and most resistant to vancomycin . The species that was most sensitive to vancomycin was acidophilus . The overall mortality rate was nearly 30% . There was a significant association between mortality and polymicrobial infection (P=0.004) . In the subset of patients with bacteremia, increased mortality was associated with inadequate treatment (P=0.001) and polymicrobial bacteremia (P=0.044). Appl Microbiol Biotechnol, 2005 Jan, 66(4), 414 - 21 Epub 2004 Jul 23. Quantification by real-time PCR of Lactococcus lactis subsp . cremoris in milk fermented by a mixed culture; Grattepanche F et al.; During cheese making, interactions between different strains of lactic acid bacteria play an important role . However, few methods are available to specifically determine each bacterial population in mixed cultures, in particular for strains of the same species . The aim of this study was to develop a real-time PCR quantification method to monitor the population of Lactococcus cremoris ATCC 19257 in mixed culture with Lactobacillus rhamnosus RW-9595M and the bacteriocin-producing microorganism Lc . diacetylactis UL719 . The specificity of the two primers 68FCa33 and 16SR308 used to amplify a 240-bp fragment of DNA from Lc . cremoris was demonstrated by conventional PCR . Using these primers for real-time PCR, the detection limit was 2 cfu/reaction or 200 cfu of Lc . cremoris ATCC 19257 per millilitre of mixed culture in milk . In pure culture batch fermentation, good correlation was obtained between real-time PCR and the conventional plating method for monitoring Lc . cremoris growth . In mixed culture batch fermentation, Lb . rhamnosus and Lc . cremoris decreased due to nisin Z production by Lc . diacetylactis . The decrease of the Lc . cremoris cell population detected by real-time PCR was not possible to observe by the plate count method in the presence of a Lc . diacetylactis population that was 1 log higher. Biotechnol Bioeng, 2005 Jan 20, 89(2), 129 - 37 Fractionation of cell mixtures using acoustic and laminar flow fields; Kumar M et al.; A fractionation method applicable to different populations of cells in a suspension is reported . The separation was accomplished by subjecting the suspension to a resonant ultrasonic field and a laminar flow field propagating in orthogonal directions within a thin, rectangular chamber . Steady, laminar flow transports the cell suspension along the chamber, while the ultrasonic field causes the suspended cells to migrate to the mid-plane of the chamber at rates related to their size and physical properties . A thin flow splitter positioned near the outlet divides the effluent cell suspension into two product streams, thereby allowing cells that respond faster to the acoustic field to be separated from those cells that respond more slowly . Modeling of the trajectories of individual cells through the chamber shows that by altering the strength of the flow relative to that of the acoustic field, the desired fractionation can be controlled . Proof-of-concept experiments were performed using hybridoma cells and Lactobacillus rhamnosus cells . The two populations of cells could be effectively separated using this technique, resulting in hybridoma/Lactobacillus ratios in the left and right product streams, normalized to the feed ratio, of 6.9 +/- 1.8 and 0.39 +/- 0.01 (vol/vol), respectively . The acoustic method is fast, efficient, and could be operated continuously with a high degree of selectivity and yield and with low power consumption . (c) 2004 Wiley Periodicals, Inc. J Infect Dis, 2005 Jan 1, 191(1), 25 - 32 Epub 2004 Dec 02. Female genital-tract HIV load correlates inversely with Lactobacillus species but positively with bacterial vaginosis and Mycoplasma hominis; Sha BE et al.; BACKGROUND: Bacterial vaginosis (BV) is associated with human immunodeficiency virus (HIV) acquisition . We examined the association between BV and BV-associated bacteria and expression of HIV in the female genital tract . METHODS: HIV RNA, lactobacilli, Gardnerella vaginalis, and Mycoplasma hominis in cervicovaginal lavage (CVL) samples were quantified by polymerase chain reaction (PCR) . Gynecologic evaluation included Nugent score assessment, Amsel criteria assessment, detection of other genital-tract infections, and dysplasia grading . CD4 cell count, plasma HIV RNA level, and antiretroviral history were obtained . RESULTS: A total of 203 CVL samples from women with Nugent scores of 7-10 (BV group) and 203 samples from women with Nugent scores of 0-3 (no-BV group) were matched by plasma HIV RNA level and analyzed . After controlling for plasma HIV RNA level and Nugent score in univariate analyses, we found that G . vaginalis and M . hominis bacterial counts, Candida vaginitis, and herpes simplex virus (HSV) were positively associated with CVL HIV RNA levels . In multivariate analysis, only lactobacilli bacterial counts (P=.006; inverse association), M . hominis bacterial counts (P=.0001; positive association), Candida vaginitis (P=.007), and HSV (P=.03) were significantly associated with CVL HIV RNA levels . CONCLUSION: Bacteria associated with BV increase genital-tract HIV RNA levels . Quantitative bacterial counts for lactobacilli and M . hominis are better correlates of CVL HIV RNA than are Nugent score or Amsel criteria . Since plasma virus and CD4 cell levels did not differ between the BV and no-BV groups, these data suggest that the bacterial flora associated with BV influence genital-tract HIV shedding. Caries Res, 2005 Jan-Feb, 39(1), 41 - 7 Caries prevalence measured with QLF after treatment with fixed orthodontic appliances: influencing factors; Boersma JG et al.; Caries prevalence on the buccal surfaces of teeth in orthodontic patients was determined with QLF and visual examination immediately after removal of fixed appliances . The number of lesions found by QLF far outnumbered that found by visual examination, but the distribution pattern was similar . 97% of all subjects and on average 30% of the buccal surfaces in a person were affected . On average, in males 40% of surfaces and in females 22% showed white spots (p < 0.01) . Caries prevalence was lower (p < 0.01) in incisors and cuspids than in molars and premolars . A positive correlation with caries prevalence was found for the bleeding scores 6 weeks after debonding and lactobacillus counts before debonding . Mutans streptococci counts, age, treatment duration, socioeconomic status and dietary habits showed no correlation with caries prevalence . J Dairy Sci, 2005 Jan, 88(1), 55 - 66 Acid and bile tolerance and cholesterol removal ability of lactobacilli strains; Liong MT et al.; Eleven strains of lactobacilli were studied for their acid and bile tolerance . Possible mechanisms of cholesterol removal by strains of lactobacilli were examined . Cholesterol assimilation as determined by the difference in cholesterol content in the medium before and after the incubation period showed that all lactobacilli strains were able to assimilate cholesterol at varying levels ranging from 12.03 to 32.25 mug/mL . Cholesterol removal was associated with growth of cultures . Binding of cholesterol to lactobacilli cells was determined using growing, heat-killed, and resting cells in phosphate buffer . Cholesterol removed by dead and resting cells ranged from 0.79 to 3.82 mg/g of dry weight compared with growing cells, which ranged from 4.53 to 16.03 mg/g of dry weight . Fatty acid methyl esters, as quantified using gas chromatography, showed changes in lipid profiles in cells grown in the presence of cholesterol compared with those grown without cholesterol . Fatty acid profiles, especially of hexadecanoic, octadecanoic, total saturated, and unsaturated acids suggested that cholesterol from the medium was incorporated into the cellular membrane . These findings suggest that strains of lactobacilli could remove cholesterol via various mechanisms and may be promising candidates for use as a dietary adjunct to lower serum cholesterol in vivo. J Dairy Sci, 2005 Jan, 88(1), 21 - 9 Fermentation pH and Temperature Influence the Cryotolerance of Lactobacillus acidophilus RD758; Wang Y et al.; The effects of 3 fermentation temperatures (30, 37, and 42 degrees C) and 3 fermentation pH (4.5, 5, and 6) on the cryotolerance of Lactobacillus acidophilus RD758 were studied in relation to their fatty acid composition . Cryotolerance was defined as the ability of the cells to recover their acidification activity after freezing and frozen storage at -20 degrees C . Better cryotolerance was obtained in cells grown at 30 degrees C or at pH 5; these cells showed no loss in acidification activity during freezing and a low rate of loss in acidification activity during frozen storage . On the other hand, cells grown at 42 degrees C or at pH 4.5 displayed poor cryotolerance . The membrane fatty acid composition was analyzed and related to the cryotolerance using principal component analysis . The improved cryotolerance observed during the freezing step was associated with a high ratio of unsaturated to saturated fatty acids, a low C18:0 content, and high C16:0 and cyclic C19:0 relative concentrations . High resistance during frozen storage was related to a high cycC19:0 concentration . Finally, the low cryotolerance observed after fermentation at pH 4.5 was explained by a low C18:2 content. Am J Clin Nutr, 2004 Dec, 80(6), 1618 - 25 Lactobacillus rhamnosus induces peripheral hyporesponsiveness in stimulated CD4+ T cells via modulation of dendritic cell function; Braat H et al.; BACKGROUND: Although it is widely recognized that the intake of so-called probiotic microorganisms is beneficial in chronic mucosal inflammation and topical allergic disease, the immunologic details explaining how such bacteria can exert these effects remain obscure . OBJECTIVE: We determined whether Lactobacillus rhamnosus can modulate T cell responses in vitro and in vivo . DESIGN: In vitro, human monocyte-derived dendritic cells (DCs) matured in the presence of L . rhamnosus were used to instruct naive CD4+ T cells; subsequently, the T cell response was assessed with the use of CD3/CD28 and interleukin (IL) 2 . Cytokine production by ex vivo-stimulated naive cells and memory T cells was measured before and after oral supplementation with L . rhamnosus in 6 healthy volunteers and 6 patients with Crohn disease . RESULTS: A decreased T cell proliferation and cytokine production, especially of IL-2, IL-4, and IL-10, was observed in CD3/CD28-stimulated T cells derived from L . rhamnosus-matured DCs . This T cell hyporesponsiveness was associated with enhanced DC-T cell interaction and normal responsiveness of T cells for IL-2 . In vivo oral supplementation of L . rhamnosus for 2 wk induced a similar T cell hyporesponsiveness, including impaired ex vivo T helper subsets 1 and 2 responses without up-regulation of immunoregulatory cytokines in cohorts of both healthy volunteers and patients with Crohn disease . CONCLUSIONS: We propose that L . rhamnosus modulates DC function to induce a novel form of T cell hyporesponsiveness; this mechanism might be an explanation for the observed beneficial effects of probiotic treatment in clinical disease. Int J Food Microbiol, 2005 Jan 1, 97(3), 307 - 15 Water activity affects heat resistance of microorganisms in food powders; Laroche C et al.; To study the factors and mechanisms involved in microorganisms' death or resistance to temperature in low-water-activity environments, a previous work dealt with the viability of dried microorganisms immobilized in thin-layer on glass beads . This work is intended to check the efficiency of a rapid heating-cooling treatment to destroy microorganisms that were dried after mixing with wheat flour or skim milk . The thermoresistance of the yeast Saccharomyces cerevisiae and the bacterium Lactobacillus plantarum were studied . Heat stress was applied at two temperatures (150 or 200 degrees C) for treatments of one of four durations (5, 10, 20, or 30 s) and at seven levels of initial water activity (a(w)) in the range 0.10 to 0.70 . This new treatment achieved a microbial destruction of eight log reductions . A specific initial water activity was defined for each strain at which it was most resistant to heat treatments . On wheat flour, this initial a(w) value was in the range 0.30-0.50, with maximal viability value at a(w)=0.35 for L . plantarum, whatever the temperature studied, and 0.40 for S . cerevisiae . For skim milk, a variation in microbial viability was observed, with optimal resistance in the range 0.30-0.50 for S . cerevisiae and 0.20-0.50 for L . plantarum, with minimal destruction at a(w)=0.30 whatever the heating temperature is. Int J Food Microbiol, 2005 Jan 1, 97(3), 285 - 96 Characterization and identification of lactic acid bacteria in "morcilla de Burgos"; Santos EM et al.; A total of 176 lactic acid bacteria (LAB) isolated from a typical Spanish blood sausage called "morcilla de Burgos" were identified by means of phenotypic characteristics and 16S rDNA RFLP (ribotyping) . LAB were isolated from "morcilla" of different producers and in different storage periods, which includes unpackaged, vacuum and modified atmosphere packaged "morcilla" and vacuum packed and pasteurised "morcilla" . The knowledge of specific spoilage bacteria of "morcilla de Burgos" will be useful to design new preservation methods to extend the shelf-life of this product . Identification made according to phenotypic and biochemical characteristics shows the majority of the isolates were heterofermentative LAB (93.2%) and eight different bacterial groups could be distinguished (A-G) . Weisella viridescens was the main species detected (42%) . In addition, Leuconostoc spp . (23.9%), Weissella confusa (11.4%) and Lactobacillus fructosus (5.7%) species were found . Few strains were phenotypically missidentified as Lactobacillus sanfrancisco, Pediococcus spp., Lactobacillus sakei/curvatus and Carnobacterium spp . and 11 strains remained unknown . Most of the leuconostocs were identified as Leuconostoc mesenteroides and Leuconostoc carnosum species . Ribotyping shows a quite good correlation with phenotypic methods, although it has been possible to identify 15 different clusters . W . viridescens and leuconostocs were also the predominant LAB . Strains identified as W . confusa by phenotypic characteristics were resolved in W . confusa and Weissella cibaria by ribotyping . Neither Carnobacterium piscicola nor Lb . sanfrancisco were identified by means of genotypic method . All Lb . fructosus strains and some more included in different phenotypic groups (17 strains in total) could not be associated with any reference strain (cluster VII). J Fam Pract . 2004 Dec;53(12):952. Lactobacillus does not prevent post-antibiotic vaginitis; Solution structure and thermolysis of Cobeta-5'-deoxyadenosylimidazolylcobamide et al.; Department of Chemistry and Biochemistry, Ohio University, Athens, Ohio 45701, USA . brownk3@ohiou.edu The solution structure of Cobeta-5'-deoxyadenosylimidazolylcobamide, Ado(Im)Cbl, the coenzyme B(12) analogue in which the axial 5,6-dimethylbenzimidazole (Bzm) ligand is replaced by imidazole, has been determined by NMR-restrained molecular modeling . A two-state model, in which a conformation with the adenosyl moiety over the southern quadrant of the corrin and a conformation with the adenosyl ligand over the eastern quadrant of the corrin are both populated at room temperature, was required by the nOe data . A rotation profile and molecular dynamics simulations suggest that the eastern conformation is the more stable, in contrast to AdoCbl itself in which the southern conformation is preferred . Consensus structures of the two conformers show that the axial Co-N bond is slightly shorter and the corrin ring is less folded in Ado(Im)Cbl than in AdoCbl . A study of the thermolysis of Ado(Im)Cbl in aqueous solution (50-125 degrees C) revealed competing homolytic and heterolytic pathways as for AdoCbl but with heterolysis being 9-fold faster and homolysis being 3-fold slower at 100 degrees C than for AdoCbl . Determination of the pK(a)'s for the Ado(Im)Cbl base-on/base-off reaction and for the detached imidazole ribonucleoside as a function of temperature permitted correction of the homolysis and heterolysis rate constants for the temperature-dependent presence of the base-off species of Ado(Im)Cbl . Activation analysis of the resulting rate constants for the base-on species show that the entropy of activation for Ado(Im)Cbl homolysis (13.7 +/- 0.9 cal mol(-1) K(-1)) is identical with that of AdoCbl (13.5 +/- 0.7 cal mol(-1) K(-1)) but that the enthalpy of activation (34.8 kcal mol(-1)) is 1.0 +/- 0.4 kcal mol(-1) larger . The opposite effect is seen for heterolysis, where the enthalpies of activation are identical but the entropy of activation is 5 +/- 1 cal mol(-1) K(-1) less negative for Ado(Im)Cbl . Extrapolation to 37 degrees C provides a rate constant for Ado(Im)Cbl homolysis of 2.1 x 10(-9) s(-1), 4.3-fold smaller than for AdoCbl . Combined with earlier results for the enzyme-induced homolysis of Ado(Im)Cbl by the ribonucleoside triphosphate reductase from Lactobacillus leichmannii, the catalytic efficiency of the enzyme for homolysis of Ado(Im)Cbl at 37 degrees C can be calculated to be 4.0 x 10(8), 3.8-fold, or 0.8 kcal mol(-1), smaller than for AdoCbl . Thus, the bulky Bzm ligand makes at best a <1 kcal mol(-1) contribution to the enzymatic activation of coenzyme B(12). Clin Infect Dis, 2004 Dec 15, 39(12), 1858 - 60 Epub 2004 Dec 15. Phenotypic differences between commercial Lactobacillus rhamnosus GG and L . rhamnosus strains recovered from blood; Ouwehand AC et al.; The isolation of clinical isolates that are indistinguishable, by molecular methods, from the probiotic strain of Lactobacillus rhamnosus GG has been reported . We compared the virulence potential of these clinical isolates with that of the probiotic strain . It was observed that all isolates differed significantly in 1 or more phenotypic properties. Eur J Nutr . 2004 Dec 1; {Epub ahead of print} Probiotic bacteria stimulate virus-specific neutralizing antibodies following a booster polio vaccination; de Vrese M et al.; BACKGROUND . Orally ingested probiotic bacteria may modulate the immune response and increase antibody titers against enteric infections by bacteria or viruses . Even though positive effects of probiotics on respiratory tract infections have been reported, overall only few studies have examined effects on virus infections concerning organs other than the gastrointestinal tract . AIM OF THE STUDY . It was the aim of the study to investigate whether and how probiotics affect the immune response to a standardized enterovirus challenge (polio) and infections not limited to the gastrointestinal tract in healthy adults . METHODS . In a randomized, controlled and double-blind study 64 volunteers consumed for 5 weeks chemically acidified clotted milk without bacteria or with 10(10)/serving (Lactobacillus rhamnosus) GG or Lactobacillus acidophilus CRL431 added . In the second week subjects were vaccinated orally against polio 1, 2 and 3 . Polio virus neutralizing serum activity, the primary parameter, was determined by the standard neutralization test (WHO) before and three times after vaccination . Polio-specific IgA, IgG and IgM were detected by ELISAs . RESULTS . Probiotics increased poliovirus neutralizing antibody titers (NT) and affected the formation of poliovirus-specific IgA and IgG in serum . The maximum increase after immunization was about 2, 2.2, or 4-fold higher, respectively, for NT, IgG or, IgA, in volunteers consuming probiotics instead of placebo . No consistent difference was noted between bacterial strains . CONCLUSIONS . Probiotics induce an immunologic response that may provide enhanced systemic protection of cells from virus infections by increasing production of virus neutralizing antibodies. J Pharm Pharm Sci, 2004 Sep 24, 7(3), 315 - 24 Artificial cell microcapsule for oral delivery of live bacterial cells for therapy: design, preparation, and in-vitro characterization; Ouyang W et al.; PURPOSE: Bacterial cells can be engineered to synthesize a wide array of disease modifying substrates such as cytokines, vaccines and antibodies; however, their use as an orally delivered therapeutic is limited by poor gastrointestinal (GI) survival and instigation of immunogenic response . Artificial cell microcapsules have been well studied as a means to overcome such problems, however, presently obtainable microcapsules have limitations . This study summarizes a novel microcapsule design specifying its preparation and GI stability in-vitro . METHOD: Multilayer APPPA microcapsules were designed, prepared and characterized in-vitro for bacterial cell oral delivery using Lactobacillus reuteri cells as a model . Microcapsule structural integrity, mechanical stability, and GI survival studies were performed in simulated gastric (SGF) and intestinal (SIF) fluids in various pH conditions at 37.2 degrees C and compared with presently available alginate/poly-l-lysine/alginate (APA) microcapsules . HPLC was used for the microcapsule membrane permeability study . RESULTS: Results show that APPPA microcapsules can be prepared for bacterial cell encapsulation and are stable in simulated GI conditions . No microcapsule damage was reported when exposed to SGF and SIF for 12 hours at 250 rpm mechanical shaking at 37.2 degrees C . In addition, 93.2+/-2.3% and 98.9+/-0.6% of microcapsules were undamaged after 24 hours in SGF and SIF respectively . Microcapsule pH stability results show that 92.8+/-3.1% of microcapsules remained intact at pH 1, 3, 5, and 7 and no damage was observed at pH 9.0 when challenged for 24 hours . When exposed for 3 hours with 250 rpm shaking at 37.2 degrees C, no damage of the microcapsules in SGF and SIF at pH, 1,3,5,7, and 9 was observed . Compared to APA microcapsules, APPPA membranes showed superior GI stability and permeability for cell encapsulation . CONCLUSION: Novel APPPA microcapsules have superior features for oral delivery of live bacterial cells and they can be used for various clinical applications . However, further study such as membrane permeability, cytotoxicity, immune protection capacity, and suitability for live bacterial cell oral delivery in-vivo is required. Appl Environ Microbiol, 2004 Dec, 70(12), 7260 - 8 Evidence for involvement of at least six proteins in adaptation of Lactobacillus sakei to cold temperatures and addition of NaCl; Marceau A et al.; Lactobacillus sakei is a lactic acid bacterium widely represented in the natural flora of fresh meat . The aim of this study was to analyze the differences in protein expression during environmental changes encountered during technological processes in which L . sakei is involved in order to gain insight into the ability of this species to grow and survive in such environments . Using two-dimensional electrophoresis, we observed significant variation of a set of 21 proteins in cells grown at 4 degrees C or in the presence of 4% NaCl . Six proteins could be identified by determination of their N-terminal sequences, and the corresponding gene clusters were studied . Two proteins belong to carbon metabolic pathways, and four can be clustered as general stress proteins . A phenotype was observed at low temperature for five of the six mutants constructed for these genes . The survival of four mutants during stationary phase at 4 degrees C was affected, and surprisingly, one mutant showed enhanced survival during stationary phase at low temperatures. Appl Environ Microbiol, 2004 Dec, 70(12), 7200 - 9 Bile-mediated aminoglycoside sensitivity in Lactobacillus species likely results from increased membrane permeability attributable to cholic acid; Elkins CA et al.; Few studies have been conducted on antimicrobial resistance in lactobacilli, presumably because of their nonpathogenic nature as anaerobic commensals . We assessed resistance in 43 type strains and isolates representing 14 species by using agar disk diffusion and MIC analysis in MRS medium . Most noteworthy were two general phenotypes displayed by nearly every strain tested: (i) they were more susceptible (up to 256-fold in some cases) to the deconjugated bile acid cholic acid than to the conjugate taurocholic or taurodeoxycholic acid, and (ii) they became susceptible to aminoglycosides when assayed on agar medium containing 0.5% fractionated bovine bile (ox gall) . Two-dimensional MIC analyses of one representative strain, Lactobacillus plantarum WCFS1, at increasing concentrations of ox gall (0 to 30.3 mg/ml) displayed corresponding decreases in resistance to all of the aminoglycosides tested and ethidium bromide . This effect was clinically relevant, with the gentamicin MIC decreasing from >1,000 to 4 mug/ml in just 3.8 mg of ox gall per ml . In uptake studies at pH 6.5, {G-3H}gentamicin accumulation increased over control levels when cells of this strain were exposed to bile acids or reserpine but not when they were exposed to carbonyl cyanide m-chlorophenylhydrazone . The effect was dramatic, particularly with cholic acid, increasing up to 18-fold, whereas only modest increases, 3- and 5-fold, could be achieved with taurocholic acid and ox gall, respectively . Since L . plantarum, particularly strain WCFS1, is known to encode bile salt hydrolase (deconjugation) activity, our data indicate that mainly cholic acid, but not taurocholic acid, effectively permeabilizes the membrane to aminoglycosides . However, at pHs approaching neutral conditions in the intestinal lumen, aminoglycoside resistance due to membrane impermeability may be complemented by a potential efflux mechanism. Appl Environ Microbiol, 2004 Dec, 70(12), 6992 - 7 Initiation of protein synthesis by a labeled derivative of the Lactobacillus casei DN-114 001 strain during transit from the stomach to the cecum in mice harboring human microbiota; Oozeer R et al.; Although studies on the survival of bacteria in the digestive tract have been reported in the literature, little data are available on the physiological adaptation of probiotics to the digestive environment . In previous work, a transcriptional fusion system (i.e., luciferase genes under the control of a deregulated promoter) was used to demonstrate that a derivative of the Lactobacillus casei DN-114 001 strain, ingested in a fermented milk and thus exhibiting initially a very weak metabolic activity, synthesized proteins de novo after its transit in the digestive tract of mice harboring human microbiota (known as human-microbiota-associated mice) . With the same genetic system and animal model, we here investigate for the first time the ability of L . casei to reinitiate synthesis in the different digestive tract compartments . In this study, most ingested L . casei cells transited from the stomach to the duodenum-jejunum within 1 h postingestion . No luciferase activity was observed in these digestive tract compartments after the first hour . At later times, the bulk of bacteria had transited to the ileum and the cecum . Luciferase synthesis was detected between 1.5 and 2.0 h postingestion at the ileal level and from 1.5 h to at least 6.0 h postingestion in the cecum, where the activity remained at a maximum level . These results demonstrate that ingested L . casei (derivative of the DN-114 001 strain) administered via a fermented milk has already reinitiated protein synthesis when it reaches the ileal and cecal compartments. Med Sci Monit . 2004 Nov 24;10(12):CR662-666 {Epub ahead of print} Effect of Lactobacillus casei supplementation on the effectiveness and tolerability of a new second-line 10-day quadruple therapy after failure of a first attempt to cure Helicobacter pylori infection; Tursi A et al.; Background: Probiotics have never been used as second-line treatment in patients resistant to a first course of anti-H . pylori treatment . Material/Methods: 70 consecutive patients with persistent H . pylori infection were enrolled and treated with ranitidine bismuth citrate (RBC) 400 mg b.d, esomeprazole or pantoprazole 40 mg/day, amoxycillin 1 g t.d, tinidazole 500 mg b.d . with (group A) or without (group B) supplementation with 750 mg daily containing 16 billion bacteria Lactobacillus casei subsp . casei DG . Esomeprazole or pantoprazole 40 mg/day was administered for a further 4 weeks in cases of active peptic ulcer or severe gastritis detected at endoscopy . In these cases endoscopy was repeated one month after conclusion of therapy . The remaining patients were checked by alC-urea breath test . Results: Sixty-six patients completed the study, 34 in group A and 32 in group B . One group A patient (2.85%) was excluded for protocol violation and one group B patient (2.85%) was lost to follow-up . 33/34 group A patients were H . pylori-negative {per-protocol: 97.05%, on intention-to-treat: 94.28%} . 5/34 patients (14.7%) showed side-effects, but all of them completed the treatment . In group B, two patients (5.71%) showed severe side-effects and were withdrawn from the study . 30/32 patients were H . pylori- negative {per-protocol: 93.75%, on intention-to-treat: 85.71% (p=n.s.)} . 11/32 patients (34.37%) showed side-effects, but all of them completed the study (p<0.05) . Conclusions: This 10-day quadruple therapy obtains a high eradication rate, but probiotic supplementation reduces side-effects and permits a slight improvement in eradicating H . pylori. Expert Rev Anti Infect Ther, 2004 Dec, 2(6), 913 - 22 Evolving issues in understanding and treating bacterial vaginosis; Marrazzo JM; Bacterial vaginosis is a synergistic polymicrobial syndrome characterized by depletion of Lactobacillus spp., especially those that produce hydrogen peroxide, and an intense increase in the quantity of commensal vaginal anaerobic bacteria to 100- to 1000-fold above normal levels . While the bacterial spectrum of these organisms has long been known to include Gardnerella vaginalis, Prevotella spp., anaerobic Gram-positive cocci, Mobiluncus spp . and Mycoplasma hominis, innovative use of molecular diagnostics has identified novel species apparently associated with this syndrome, including Atopobium vaginalis . Effecting resolution of bacterial vaginosis is important, in particular for the 8 to 23% of women afflicted with symptomatic disease during their reproductive years . Bacterial vaginosis has been consistently associated with numerous adverse sequelae related to the upper genital tract, including pelvic inflammatory disease and postsurgical infection in the setting of invasive gynecologic procedures, and may increase women's risk of acquiring HIV infection . Pregnant women with bacterial vaginosis experience a higher rate of preterm delivery and low-birth-weight infants . While antibiotics with activity against anaerobes--typically, metronidazole and clindamycin applied vaginally or taken orally--are the mainstays of therapy, bacterial vaginosis frequently recurs . For these reasons, innovative approaches to therapy are urgently required. J Chemother, 2004 Oct, 16(5), 463 - 7 Microbiological evaluation of commercial probiotic products available in Italy; Drago L et al.; Scientific evidence of the prevention and therapy of some intestinal diseases is accumulating in regard to probiotic products . However, sufficient information on the use of probiotics in specific therapies is not yet available and, above all, there is no clear legislation about these products in Europe . In this study, we evaluated five different probiotic products commercially available in Italy for their qualitative and quantitative microbial content after about 12 and 22 months of storage . We also evaluated the stability of lactobacilli to 0.3% bile salts and to pH of 3.58 and 7.98 . There were discrepancies between the declared content and our results found after storage for 4 of the tested products . Bile salts and basic pH did not affect the growth of the lactobacilli tested, while for 2 tested products 6 hours at acid pH produced a complete inhibition of bacterial growth . Our results suggest the need for clear legislation and adequate control of the manufacturing of probiotic products. Eur J Dermatol, 2004 Nov-Dec, 14(6), 400 - 6 Clinical usefulness of oral itraconazole, an antimycotic drug, for refractory atopic dermatitis; Ikezawa Z et al.; We investigated the clinical usefulness of oral itraconazole for refractory atopic dermatitis in a crossover study . Patients with refractory atopic dermatitis were divided into two groups: Group A; a combination of itraconazole plus a conventional lactobacillus preparation was administered for 8 weeks, followed by lactobacillus preparation alone for 8 weeks, Group B; lactobacillus preparation alone was administered for 8 weeks, followed by itraconazole plus lactobacillus for 8 weeks . In both groups, a decrease in dose or strength of concomitant topical steroids was observed at the end of the treatment course of itraconazole, and improvement of parameters such as eosinophil count, serum IgE level and specific IgE antibody titers to fungi was also observed after the adm