Growth-inhibitory effects of Galla Rhois-derived tannins on intestinal bacteria

The growth-inhibitory activity of Galla Rhois-derived materials towards 17 intestinal bacteria was evaluated using an impregnated paper disc method. The biologically active components of Galla Rhois were characterized as the tannins methyl gallate (MG) and gallic acid (GA) by spectral analysis. The growth responses varied with bacterial strain tested. In the test using 10 mg disc⁻¹, MG and GA produced a clear inhibitory effect on harmful bacteria such as Clostridium perfringens , Cl. paraputrificum , Eubacterium limosum , Bacteroides fragilis , Staphylococcus aureus and Escherichia coli . Methyl gallate showed no growth-inhibitory activity towards Bifidobacterium adolescentis or B. longum whereas the growth of B. bifidum , B. breve , B. infantis , B. animalis , B. thermophilum , Lactobacillus acidophilus , Lact. plantarum and Streptococcus faecalis was slightly affected. However, GA did not adversely affect the growth of the bifidobacteria and lactobacilli. At 5 mg disc⁻¹, MG significantly inhibited the growth of Cl. perfringens and Cl. paraputrificum but did not affect the growth of the bifidobacteria and lactobacilli. At 1 mg disc⁻¹, MG greatly inhibited the growth of Cl. perfringens alone. These results may be an indication of at least one of the pharmacological actions of Galla Rhois.     

莳萝蒿精油成分分析及抑菌活性机理探究

为了确定莳萝蒿精油的化学成分,并探究其抑菌活性及抑菌机理.该研究采用水蒸气蒸馏法提取莳萝蒿精油,并通过气相色谱-质谱联用法测定其化学成分.采用抑菌圈法、二倍稀释法和生长曲线法测定精油的抑菌活性,采用电导率法和扫描电镜法探究精油的抑菌机理.结果表明:(1)莳萝蒿精油的主要化学成分包括醇类(47.12％)和萜烯类(19.9...     

固定化细胞利用离子交换树脂萃取发酵乳酸的研究

本文提出了利用海藻酸钙凝胶包埋固定化乳酸菌生产乳酸,用离子交换树脂从发酵液中分离出乳酸的新方法。该法成功地消除了产物乳酸对乳酸菌生长和产物乳酸形成的抑制作用,使发酵时间由１２０小时缩短到９６小时,乳酸的体积生产率由０．３２８ｇ／Ｌ·ｈ提高到０．４３２ｇ／Ｌ·ｈ。     

Effects of Rosa rugosa petals on intestinal bacteria

The effects of pulverized petal of Rosa rugosa on the growth of 10 species of intestinal and pathogenic bacteria were investigated. Growth of bifidobacteria and lactobacilli was not affected by the addition of the petal in plate cultivation. However, the growth of Bacteroides vulgatus, Escherichia coli, Staphylococcus aureus, and Bacillus cereus was completely inhibited by the addition of 0.1, 0.5, 0.1, and 0.05% (w/v) of the petal respectively. In liquid cultivation, the addition of the petal (0.5%) stimulated the growth of Bifidobacterium breve and slightly inhibited the growth of Lactobacillus salivarius. But the growth of E. coli, S. aureus, B. cereus, and Salmonella sp. was inhibited by nearly 50%. Hydrolyzable tannins isolated from R. rugosa, rugosin D, and tellimagradin II showed antibacterial activities against E. coli, S. aureus, B. cereus, and Salmonella sp., but little or no effect against Bif. breve and L. salivarius. R. rugosa petal showed selective antibacterial activities against intestinal and pathogenic bacteria, and the selectivity resembled that of prebiotics such as oligosaccharides and dietary fiber. Hydrolyzable tannins in R. rugosa, such as rugosin D and tellimagradin II, must be active constituents.     

Seasonal variation in allelopathic potential of<Emphasis Type="Italic">Artemisia princeps</Emphasis> var.<Emphasis Type="Italic">orientalis</Emphasis> on plants and microbes

We investigated seasonal variations in allelopathic potential ofArtemisia princeps var.orientalis. Aqueous and meth-anol extracts and volatile substances were prepared in the laboratory from samples collected monthly (April through October). Their impacts were then assessed on the germination and seedling growth ofLactuca sativa andAchyranthes japonica. The allelopathic potential varied with the time of sample collection and the concentration tested. For example, germination ofL. sativa was not inhibited by the aqueous extract but seedling growth (shoots and roots) was, with its seasonal effect being significant. ForA. japonica, seed germination was not inhibited at lower concentrations (except for August samples). However, at higher concentrations and in certain months (especially July), germination was more negatively affected. The degree of seedling growth inhibition also differed by month and by extract concentration, with roots being impacted more than shoots. Volatile substances also had a time-dependent influence on the germination and seedling elongation ofA. japonica. In a separate experiment, the ethyl-acetate and water fractions of a crude methanol extract were prepared monthly fromA. princeps var.orientalis. Here, we examined their antimicrobial activities against three gram-positive bacteria (Bacillus cereus, Bacillus subtilis, andStaphylococcus aureus), two gramnegative bacteria (Escherichia coli andPseudomonas fluorescens), and one lactic acid bacterium,Lactobacillus plantar urn. The ethyl-acetate fraction that was sampled in September was remarkably potent againstB. cereus andB. subtilis, whereas the water fraction collected in August and September showed great antimicrobial activity against the grampositive and -negative bacteria. In contrast,L. plantarum was not inhibited by the water fraction, regardless of the sampling month. Likewise, the ethyl-acetate and water fractions collected in April and October had the lowest levels of antimicrobial activity.     

A method for the selective enumeration and isolation of ruminal Lactobacillus and Streptococcus

Ruminal lactic acid-producing bacteria were selectively isolated and enumerated using a one hour aerobic exposure prior to incubation on a semi-selective Lactobacillus medium, MRS, under anaerobic conditions. The technique allowed growth of pure cultures of ruminal Lactobacillus spp. and Streptococcus bovis without supporting the growth of pure cultures of any of the prominent ruminal bacterial species. In mixed cultures, the one hour aerobic pre-incubation inhibited the growth of the obligate anaerobic ruminal bacteria which can otherwise grow on the MRS medium, and the subsequent anaerobic incubation permitted maximal recovery of the weakly aerotolerant ruminal lactic acid-producing Lactobacillus spp. and Streptococcus spp. The efficacy of this technique in selecting exclusively for the lactic acid-producing bacteria was also demonstrated from populations of rumen bacteria from mixed culture end-point in vitro fermentation, continuous in vitro culture and isolations from fresh ruminal samples.     

Brachyspira (Serpulina) pilosicoli of human origin interfere with the growth of Clostridium perfringens alpha-toxin producer

Brachyspira (Serpulina) pilosicoli of human origin interfere with the growth of Clostridium perfringens alpha-toxin producer reducing the clostridial growth area and colonies number when bacteria were cultivated together in sheep blood agar plates. The growth inhibition of C. perfringens was only observed when B. (S.) pilosicoli grew 72-96 hours sooner than C. perfringens and after the inoculum of this latter the plates were anaerobically incubated for additional 48 hours. The phenomenon was observed at concentrations of B. (S.) pilosicoli ranging from 10(7) to 10(4) CFU/ml and at concentrations of C. perfringens ranging from 10(7) to 10(1) CFU/ml when the bacteria were 0-10 mm away from each other. When B. (S.) pilosicoli and C. perfringens were inoculated at the same time and when B. (S.) pilosicoli grew 24-48 hours sooner than C. perfringens, the clostridial growth inhibition was not appreciated and only a cooperative haemolysis was observed between the bacteria.     

Behaviour of the pharmaceutical probiotic preparation VSL#3 in human ileostomy effluent containing its own natural elements

The pharmaceutical probiotic VSL#3 (300 billion cfu/g lactic acid bacteria & bifidobacteria) was inoculated into human ileostomy effluent (HIE) to assess its behaviour vs the ileo-caecal tract. Separately, yogurt bacteria (yog) and bifidobacteria (Bif) present in VSL#3 were also inoculated into HIE. During 37 degrees C incubation (anaerobic condition) at zero, six and 24 hours, both cell growth in control HIE and indigenous Bif growth in HIE+yog were observed. Cells remained viable and metabolically active as shown by the increase in L(+) lactic acid in HIE+VSL#3 and HIE+yog and the pH decreased (approx. 5.5 compared with the 6.2 of control HIE). Total SCFA Short Chain Fatty Acids decreased in HIE+yog and HIE+VSL#3 at 6h and in all cultures at 24h; butyric acid decreased in HIE+Bif and HIE+VSL#3. Generally in vivo the bacteria remain in the ileo-caecal tract no longer than six h, therefore VSL#3 bacteria seem able to pass this barrier viably, colonizing the large bowel.     

The antimicrobial activity of heyneanol A extracted from the root of Taiwanese wild grape

Aims:  To search for antimicrobial compounds against pathogenic bacteria from grape vines ( Vitis spp.). To investigate the antimicrobial efficacy of active compounds towards methicillin-resistant Staphylococcus aureus (MRSA).
Methods and Results:  The root extracts of taiwanese wild grape ( Vitis thunbergii var. taiwaniana ) showed marked activities against Gram-positive bacteria using the disc diffusion method. After purification, the active compound 1 was confirmed as heyneanol A by mass spectroscopy and nuclear magnetic resonance. Heyneanol A showed an minimum inhibitory concentration (MIC) value of 2  μ g ml⁻¹ towards MRSA and a value of 2 to 4  μ g ml⁻¹ for Enterococcus faecium , S. aureus , Streptococcus agalactiae and Streptococcus pyogenes . In addition, the contents of heyneanol A were determined as 36 mg g⁻¹ in roots of taiwanese wild grape.
Conclusions:  The root extracts of grapevines have good antimicrobial activities towards some strains of Gram-positive pathogens. Heyneanol A, the major antimicrobial compound, is especially active towards MRSA. In addition, the abundances of heyneanol A and other stilbenes in the roots of grapevines make it possible to produce natural antimicrobial compounds from this plant species.
Significance and Impact of the Study:  This study reports for the first time the antimicrobial compounds in the root extracts of grapevines. The results will have clinical significance owing to their activities against MRSA.     

Selecting for lactic acid producing and utilising bacteria in anaerobic enrichment cultures

Lactic acid-producing bacteria are important in many fermentations, such as the production of biobased plastics. Insight in the competitive advantage of lactic acid bacteria over other fermentative bacteria in a mixed culture enables ecology-based process design and can aid the development of sustainable and energy-efficient bioprocesses. Here we demonstrate the enrichment of lactic acid bacteria in a controlled sequencing batch bioreactor environment using a glucose-based medium supplemented with peptides and B vitamins. A mineral medium enrichment operated in parallel was dominated by Ethanoligenens species and fermented glucose to acetate, butyrate and hydrogen. The complex medium enrichment was populated by Lactococcus, Lactobacillus and Megasphaera species and showed a product spectrum of acetate, ethanol, propionate, butyrate and valerate. An intermediate peak of lactate was observed, showing the simultaneous production and consumption of lactate, which is of concern for lactic acid production purposes. This study underlines that the competitive advantage for lactic acid-producing bacteria primarily lies in their ability to attain a high biomass specific uptake rate of glucose, which was two times higher for the complex medium enrichment when compared to the mineral medium enrichment. The competitive advantage of lactic acid production in rich media can be explained using a resource allocation theory for microbial growth processes.     

Effect of Lactic Acid Bacteria on Growth of Staphylococcus aureus

Cultures of lactic acid bacteria, mostly from foods, were tested for their effect on the growth of Staphylococcus aureus in Trypticase Soy Broth (BBL). Some of the effectors, e.g., Streptococcus faecalis, S. faecium, Lactobacillus lactis, L. brevis, and Leuconostoc mesenteroides, stimulated growth of S. aureus during early hours of growth, especially at higher temperatures of incubation, but most cultures were inhibitory, and some (S. faecium and L. mesenteroides) were even killing by the time of attainment of the maximal phase of growth of the Staphylococcus. Low-temperature meat lactobacilli and Leuconostoc dextranicum inhibited S. aureus at 10, 15, 20, and 25 C throughout its growth. Streptococcus faecalis var. liquefaciens inhibited at these temperatures and at 30 and 37 C, as well. When the ratio of effectors to staphylococci in the inoculum was 100:1, the three enterococci, the meat Lactobacillus, and L. dextranicum prevented the attainment of 5 x 10(6) staphylococci per milliliter at 15 C, and all but the meat Lactobacillus did so at 22 C. A ratio of 1:1 accomplished similar results at 15 C, except that S. aureus was only delayed for 12 hr by S. faecalis. A ratio of 1:100 usually was ineffective. In general, the more effector bacteria there were in the inoculum, the greater was the overall inhibition (or stimulation) of S. aureus. Inhibition was most effective at 10 or 15 C, less so at 20 or 25 C, and least at 30 or 37 C, whereas stimulation during early growth was greater at the higher temperatures. Results with different strains of the effectors and with two strains of S. aureus were similar, for the most part.     

内生贝莱斯芽胞杆菌生物拮抗活性的研究

目的研究内生菌的生物拮抗活性。方法基于植物贴片法、形态学与分子生物学特征等分离、鉴定内生菌;利用纸片扩散法检测分离株拮抗病原性细菌、植物部分内生菌及耐药性细菌的生物活性。结果分离株HBB5的菌落呈乳白色、黏液性、皱褶状凸起,镜下见革兰阳性、单个或成双或短链状排列杆菌;序列分析显示,HBB5菌的16SrRNA基因序列与GenBank数据库中贝莱斯芽胞杆菌(Bacillus velezensis)同源性为100%,鉴定为B.velezensis HBB5。拮抗试验显示,B.velezensis HBB5抑制Staphylococcus aureus、Enterococcus faecalis、Escherichia coli、Salmonella typhi、Salmonella paratyphi A、Shigella dysenteriae、内生Penicillium HBF3及Klebsiella ornithinolytica B1645-1等菌株生长,其与内生B.subtilis SWB8无明显抑制现象。结论首次从盾叶薯蓣植物中分离出Bacillus velezensis,其显示了广谱的抗病原细菌活性及一定的拮抗耐药菌的能力,可能参与宿主植物疾病的防御。     

Differential Effects of Oxygen and Oxidation Reduction Potential on the Multiplication of Three Species of Anaerobic Intestinal Bacteria

The sensitivity of three strains of anaerobic intestinal bacteria, Clostridium perfringens, Bacteroides fragilis, and Peptococcus magnus, to the differential effects of oxygen and adverse oxidation-reduction potential was measured. The multiplication of the three organisms was inhibited in the presence of oxygen whether the medium was at a negative oxidation-reduction potential (Eh of -50 mV), poised by the intermittent addition of dithiothreitol, or at a positive oxidation-reduction potential (Eh of near +500 mV). However, when these organisms were cultured in the presence of oxygen, no inhibition was observed, even when the oxidation-reduction potential was maintained at an average Eh of +325 mV by the addition of potassium ferricyanide. When the cultures were aerated, the growth patterns of the three organisms demonstrated different sensitivities to oxygen. P. magnus was found to be the most sensitive. After 2 h of aerobic incubation, no viable organisms could be detected. B. fragilis was intermediately sensitive to oxygen with no viable organisms detected after 5 h of aerobic incubation. C. perfringens was the least sensitive. Under conditions of aerobic incubation, viable organisms survived for 10 h. During the experiments with Clostridium, no spores were observed by spore staining.     

奶牛阴道菌群分析与乳酸菌的分离及鉴定

目的研究奶牛阴道菌群,并从健康奶牛阴道分离出产酸能力很强的乳酸菌。方法采用常规的方法对奶牛阴道进行细菌的分离及鉴定,并进行菌群分析。结果健康奶牛阴道优势菌群主要为乳酸菌(P<0.01),屡配不孕奶牛阴道优势菌群主要为金黄色葡萄球菌(P<0.01);从健康奶牛阴道分离出的乳酸菌为55株,其中产酸能力很强的6株乳酸菌鉴定结果分别为Lactobacillus plantarum、Lactobacillus brevis、Enterococcus faecalis、Lactococcus garvieae、Lactobacillus kitasatonis和Lactobacillus amylovorus。结论奶牛阴道菌群中分离的6株乳酸菌可作为潜在的奶牛阴道微生态制剂进行深入研究。     

Construction of shuttle cloning vectors for Bacteroides fragilis and use in assaying foreign tetracycline resistance gene expression

Shuttle vectors capable of replication in both Escherichia coli and Bacteroides fragilis have been developed. Conjugal transfer of these plasmids from E. coli to B. fragilis is facilitated by inclusion of the origin of transfer of the IncP plasmid RK2. The vectors pDK1 and pDK2 provide unique sites for cloning selectable markers in Bacteroides. pOA10 is a cosmid vector containing the replication region of pCP1 necessary for maintenance in Bacteroides. pDK3, pDK4.1, and pDK4.2 contain the Bacteroides clindamycin resistance gene allowing selection and maintenance in B. fragilis of plasmids containing inserted DNA fragments. pDK3 was used to test the expression in B. fragilis of five foreign tetracycline resistance (TcR) genes. The tetA, -B, and -C markers from facultative gram-negative bacteria, as well as a TcR determinant from Clostridium perfringens, did not express TcR in B. fragilis. The tetM gene, originally described in streptococci, encoded a small but reproducible increase of TcR in Bacteroides. These studies demonstrate the utility of shuttle vectors for introducing cloned genes into Bacteroides and underscore the differences in gene expression in these anaerobes.     

Effect of Selected Lactic Acid Bacteria on Growth of Staphylococcus aureus and Production of Enterotoxin

Representative strains of 15 species of lactic acid bacteria were examined for their ability to influence growth of Staphylococcus aureus and production of enterotoxin in associative culture. Among the organisms used as effectors the streptococci were most inhibitory, followed by Pediococcus cerevisiae. The lactobacilli and Leuconostoc citrovorum were not inhibitory to growth and only slightly inhibitory to production of enterotoxin. Enterotoxin was detected in all cultures in which the population of S. aureus reached 8 x 10(7) per ml. At lower S. aureus populations no enterotoxin was detected after incubation for 48 h. Mechanisms of inhibition of growth and enterotoxin production by S. aureus strain 243 grown in association with Streptococcus lactis A64 or P. cerevisiae 10791 in APT broth were investigated. Competition for vital nutrients, especially niacin and biotin, and probably production of hydrogen peroxide contribute to inhibition. Production of lactic acid appears to inhibit growth of S. aureus in the early but not the late stages of incubation.     

Capsular polysaccharides and lipopolysaccharides from two Bacteroides fragilis reference strains: chemical and immunochemical characterization.

Fermentor growth of Bacteroides fragilis under controlled conditions in a complex medium containing 1% glucose and 10% fetal calf serum resulted in high yields of bacteria. After hot phenol-water extraction of the organisms, capsular polysaccharide was isolated from the aqueous phase and purified by Sephacryl S-300 chromatography in a buffer with 3% sodium deoxycholate. Lipopolysaccharide was isolated by phenol-chloroform-light petroleum ether extraction. The capsular polysaccharide from B. fragilis strain NCTC 9343 contained six sugars: L-fucose, D-galactose, D- and L-quinovosamine, D-glucosamine, and galacturonic acid. The capsule of strain ATCC 23745 also contained D-glucose, L-fucosamine, L-rhamnosamine, and a 3-amino-3,6-dideoxyhexose but lacked D-quinovosamine. The latter capsule also contained alanine (4%). The capsular polysaccharides were different immunochemically by ELISA inhibition. The lipopolysaccharide of both strains contained the same sugars (L-rhamnose, D-glucose, D-galactose, and D-glucosamine) and fatty acids (13-methyl-tetradecanoic and 3-hydroxy-hexadecanoic and 3-hydroxy-15 methyl-hexadecanoic as major constituents) and were identical by ELISA inhibition.     

新疆传统发酵乳品中乳酸菌与 酵母菌的益生特性

目的观察新疆传统发酵乳品中分离的14种菌株的生长特点及产酸能力,筛选出具有较强耐胆盐能力,并能在人工胃肠液中存活的菌株。方法对10株乳酸菌和4株酵母菌进行生长曲线、pH、耐胆盐能力和耐人工胃肠液检测。结果 10株乳酸菌和4株酵母菌具有良好的生长曲线和产酸能力;马乳酒样乳杆菌具有较强的耐胆盐能力;希氏乳杆菌、马乳酒样乳杆菌、乙醇假丝酵母和东方伊萨酵母具有较强的耐人工胃液能力;乳酸乳球菌、哈尔滨乳杆菌、瑞士乳杆菌、马乳酒样乳杆菌、乙醇假丝酵母和东方伊萨酵母具有较强的耐人工肠液能力。结论 10株乳酸菌和4株酵母菌具有优良的益生特性,有望成为益生菌制剂的备用菌株。     

Effect of premix ingredients and its inoculation with lactic acid-producing bacteria on ensiling of poultry droppings with conventional feeds

A premix of poultry droppings (25 parts) and wheat straw (65 parts) was inoculated with a mixed culture of Lactobacillus plantarum and Streptococcus faecalis in the presence or absence of sugarcane molasses. The inclusion of lactic acid-producing bacteria and molasses in the premix resulted in reduced pH, ammonia nitrogen and total volatile fatty acids and increased production of lactic acid during a fermentation time of 20 d at 32–35°C. The sugarcane molasses and part of the wheat straw can be replaced with green maize fodder in the vegetative stage without any adverse effect on the silage characteristics. With an increasing level of green maize in the premix, there was a decrease in pH, dry matter of premix and wastelage and a significant increase in soluble sugars of premix and titrable acidity and lactic acid content of wastelage. A good wastelage can be produced when the premix consists of poultry droppings, wheat straw and green maize in the ratio of 25: 15:60, respectively, and it is inoculated with lactic acid-producing bacteria.     

Influence of pH, nutrient availability, and growth rate on amine production by Bacteroides fragilis and Clostridium perfringens

Dimethylamine, methylamine, propylamine, and pyrrolidine were the major amines formed by Bacteroides fragilis NCDO 2217 during the active phase of growth in batch culture. Production of these metabolites was strongly pH dependent and was optimal under acidic conditions (pH 6.0). Low pH also favored the formation of pyrrolidine, cadaverine, and dimethylamine by Clostridium perfringens C523, but the reverse was the case with putrescine, butylamine, and propylamine, where production was maximal at neutral pH. B. fragilis was grown in continuous culture under either starch or casein limitation. Amine formation was influenced by carbohydrate availability and was greatest when the bacteria were grown at high growth rates (dilution rate, 0.20/h) under starch limitation, where they constituted about 18% of the total fermentation products measured. Amine production was optimal and increased concomitantly with growth rate when C. perfringens was grown in glucose-limited continuous culture. Under conditions of high growth rate and glucose limitation, amines accounted for approximately 27% of the fermentation products measured. When glucose in the feed medium was increased from 5 to 15 g/liter, amine production was repressed, and under these nutritional conditions the growth rate had little effect on the process.