内蒙古呼伦贝尔地区传统发酵乳中乳酸菌的多样性分析

【目的】对内蒙古呼伦贝尔地区传统发酵乳制品中乳酸菌资源的生物多样性进行研究。【方法】采用纯培养和16S rRNA基因序列分析法对内蒙古呼伦贝尔地区传统发酵乳中的乳酸菌进行多样性分析。【结果】从8份传统发酵乳制品(6份酸牛奶和2份酸马奶)样品中分离到24株乳酸菌,通过16S rRNA基因序列分析和系统进化关系分析将24株乳酸菌鉴定为2株Lactobacillus kefiranofaciens、2株Lactobacillus kefiri、5株Lactobacillus paracasei、3株Lactobacillus plantarum、1株Lactobacillus rhamnosus、6株Lactococcus lactis subsp.lactis、2株Leuconostoc mesenteroides subsp.dextranicum、2株Streptococcus thermophilus和1株Enterococcus faecium。【结论】Lactococcus lactis subsp.lactis为内蒙古呼伦贝尔地区传统发酵乳制品的优势菌种,占总分离株的25%,其次为Lactobacillus paracasei,占总分离株的20.83%。     

Production of lactic acid from date juice extract with free cells of single and mixed cultures of Lactobacillus casei and Lactococcus lactis

The production of lactic acid from date juice by single and mixed cultures of Lactobacillus casei and Lactococcus lactis was investigated. In the present conditions, the highest concentration of lactic acid (60.3 g l⁻¹) was obtained in the mixed culture system while in single culture fermentations of Lactobacillus casei or Lactococcus lactis, the maximum concentration of lactic acid was 53 and 46 g l⁻¹, respectively. In the case of single Lactobacillus casei or Lactococcus lactis, the total percentage of glucose and fructose utilized were 82.2; 94.4% and 93.8; 60.3%, respectively, whereas in the case of mixed culture, the total percentage of glucose and fructose were 96 and 100%, respectively. These results showed that the mixed culture system gave better results than single cultures regarding lactic acid concentration, and sugar consumption.     

Antifungal activity of strains of lactic acid bacteria isolated from a semolina ecosystem against Penicillium roqueforti, Aspergillus niger and Endomyces fibuliger contaminating bakery products

Thirty samples of Italian durum wheat semolina and whole durum wheat semolina, generally used for the production of Southern Italy's traditional breads, were subjected to microbiological analysis in order to explore their lactic acid bacteria (LAB) diversity and to find strains with antifungal activity. A total of 125 presumptive LAB isolates (Gram-positive and catalase-negative) were characterized by repetitive extragenic palindromic-PCR (REP-PCR) and sequence analysis of the 16S rRNA gene, leading to the identification of the following species: Weissella confusa, Weissella cibaria, Leuconostoc citreum, Leuconostoc mesenteroides, Lactococcus lactis, Lactobacillus rossiae and Lactobacillus plantarum. The REP-PCR results delineated 17 different patterns whose cluster analysis clearly differentiated W. cibaria from W. confusa isolates. Seventeen strains, each characterized by a different REP-PCR pattern, were screened for their antifungal properties. They were grown in a flour-based medium, comparable to a real food system, and the resulting fermentation products (FPs) were tested against fungal species generally contaminating bakery products, Aspergillus niger, Penicillium roqueforti and Endomyces fibuliger. The results of the study indicated a strong inhibitory activity – comparable to that obtained with the common preservative calcium propionate (0.3% w/v) – of ten LAB strains against the most widespread contaminant of bakery products, P. roqueforti. The screening also highlighted the unexplored antifungal activity of L. citreum, L. rossiae and W. cibaria (1 strain), which inhibited all fungal strains to the same or a higher extent compared with calcium propionate. The fermentation products of these three strains were characterized by low pH values, and a high content of lactic and acetic acids.     

Influence of ripening container on the lactic acid bacteria population in Tulum cheese

The objective of the present study was to investigate the influence of container material (plastic or goat-skin bag) on the growth of lactic acid bacteria in Tulum cheese during 9 months of ripening. The lactic acid bacteria in Tulum cheeses were periodically counted on MRS and M17 agars throughout ripening. Results showed that the highest counts of lactic acid bacteria on MRS or M17 were observed at the beginning of ripening and their counts decreased during later stages of ripening. The cheese samples ripened in plastic bags exhibited higher numbers of LAB on MRS and M-17 agars than those ripened in goat-skin bags. A total of 112 strains of lactic acid bacteria were isolated from Tulum cheeses ripened in plastic or goat-skin bags during ripening. The lactic acid bacteria present in the cheese were classified by Microbial Identification System (MIS) based on a comparison of the fatty acid methyl ester profiles. Different species including Enteroccocus, Lactobacillus, Streptococcus, Lactococcus and Pediococcus genera were found in unripened cheese. As ripening proceeded, the species Streptococcus and Lactococcus disappeared and the percentages of the species Enterococcus was unchanged in both containers. There were slight differences between the cheeses ripened in plastic or goat-skin bags in terms of the profiles of lactic acid bacteria isolated. Some species including L. brevis, L. mesenteroides subsp. dextranicum, P. damnosus and E. mundtii were isolated only in the cheeses ripened in plastic bags; however, L. coryniformis and L. malafermentans were isolated only in the cheeses ripened in goat-skin bags at 6 or 9 months of ripening. Also the numbers of E. faecalis isolates were higher in the cheeses ripened in plastic containers than cheeses ripened goat-skin bags at the 6 or 9 months of ripening. The results showed that Lactobacillus and Enterococcus were the predominant species in matured Tulum cheeses in both ripening containers. It seemed possible to produce Tulum cheese with similar characteristics from both the containers used.     

Genomic organization of lactic acid bacteria

Current knowledge of the genomes of the lactic acid bacteria, Lactococcus lactis and Streptococcus thermophilus, and members of the genera Lactobacillus, Leuconostoc, Pediococcus and Carnobacterium is reviewed. The genomes contain a chromosome within the size range of 1.8 to 3.4 Mbp. Plasmids are common in Lactococcus lactis (most strains carry 4–7 different plasmids), some of the lactobacilli and pediococci, but they are not frequently present in S. thermophilus, Lactobacillus delbrueckii subsp. bulgaricus or the intestinal lactobacilli. Five IS elements have been found in L. lactis and most strains carry multiple copies of at least two of them; some strains also carry a 68-kbp conjugative transposon. IS elements have been found in the genera Lactobacillus and Leuconostoc, but not in S. thermophilus. Prophages are also a normal component of the L. lactis genome and lysogeny is common in the lactobacilli, however it appears to be rare in S. thermophilus. Physical and genetic maps for two L. lactis subsp. lactis strains, two L. lactis subsp. cremoris strains and S. thermophilus A054 have been constructed and each reveals the presence of six rrn operons clustered in less than 40% of the chromosome. The L. lactis subsp. cremoris MG1363 map contains 115 genetic loci and the S. thermophilus map has 35. The maps indicate significant plasticity in the L. lactis subsp. cremoris chromosome in the form of a number of inversions and translocations. The cause(s) of these rearrangements is (are) not known. A number of potentially powerful genetic tools designed to analyse the L. lactis genome have been constructed in recent years. These tools enable gene inactivation, gene replacement and gene recovery experiments to be readily carried out with this organism, and potentially with other lactic acid bacteria and Gram-positive bacteria. Integration vectors based on temperate phage attB sites and the random insertion of IS elements have also been developed for L. lactis and the intestinal lactobacilli. In addition, a L. lactis sex factor that mobilizes the chromosome in a manner reminiscent to that seen with Escherichia coli Hfr strains has been discovered and characterized. With the availability of this new technology, research into the genome of the lactic acid bacteria is poised to undertake a period of extremely rapid information accrual.     

Synergism among lactic acid, sulfite, pH and ethanol in alcoholic fermentation of Saccharomyces cerevisiae (PE-2 and M-26)

Summary The industrial production of ethanol is affected mainly by contamination by lactic acid bacteria besides others factors that act synergistically like increased sulfite content, extremely low pH, high acidity, high alcoholic content, high temperature and osmotic pressure. In this research two strains of Saccharomyces cerevisiae PE-2 and M-26 were tested regarding the alcoholic fermentation potential in highly stressed conditions. These strains were subjected to values up to 200 mg NaHSO₃ l⁻¹, 6 g lactic acid l⁻¹, 9.5% (w/v) ethanol and pH 3.6 during fermentative processes. The low pH (3.6) was the major stressing factor on yeasts during the fermentation. The M-26 strain produced higher acidity than the other, with higher production of succinic acid, an important inhibitor of lactic bacteria. Both strains of yeasts showed similar performance during the fermentation, with no significant difference in cell viability.     

Preliminary characterization of wine lactobacilli able to degrade arginine

Lactobacillus strains able to degrade arginine were isolated and characterized from a typical red wine. All the strains were gram-positive, catalase-negative and produced both D- and L-lactate from glucose. Strains L2, L3, L4, and L6 were able to produce CO₂ from glucose; however, production of CO₂ from glucose was not observed in strains L1 and L5, suggesting that they belong to the homofermentative wine lactic acid bacteria (LAB) group. All of the lactobacilli were tested for their ability to ferment 49 carbohydrates. The sugar fermentation profile of strain L1 was unique, suggesting that this strain belonged to Lactococcus lactis ssp. cremoris, a non-typical wine LAB. Furthermore, a preliminary typing was performed by using a random amplified polymorphic DNA analysis (RAPD-PCR analysis).     

Microbial Diversity in Ngari,Hentak and Tungtap,Fermented Fish Products of North-East India

Ngari, hentak and tungtap are traditional fermented fish products of North-East India. Eighteen samples of ngari, hentak and tungtap were collected and were analysed for microbial load. Lactic acid bacteria, endospore-forming rods, yeasts and aerobic mesophilic counts ranged from 4.0 to 7.2, 3.3–4.6, <1–3.5 and 4.3–7.3 log c.f.u./g, respectively. Lactic acid bacteria were identified as Lactococcus lactis subsp. cremoris, Lactococcus plantarum, Enterococcus faecium, Lactobacillus fructosus, Lactobacillus amylophilus, Lactobacillus coryniformis subsp. torquens and Lactobacillus plantarum. Endospore-forming rods were identified as Bacillus subtilis and Bacillus pumilus, aerobic coccal strains were identified as Micrococcus. Yeasts were identified as species of Candida and Saccharomycopsis. Pathogenic contaminants were detected in all samples, however, none of the sample contained more than 10² c.f.u./g of Bacillus cereus, 10³ c.f.u./g of Staphylococcus aureus and enterobacteriaceae population, respectively. Enzymatic and antimicrobial activities of the isolates were tested. None of the strains produced biogenic amines in the method applied. Most strains of LAB had a high degree of hydrophobicity, indicating their ‘probiotic’ characters. This study has demonstrated the microbial diversity within the species of lactic acid bacteria, Bacillus and yeasts. This revised version was published online in July 2006 with corrections to the Cover Date.     

Cytotoxic, antioxidative, and ACE inhibiting activities of dolsan leaf mustard juice (DLMJ) treated with lactic acid bacteria

This study was performed to know whether there is any change of physiological activity in DLMJ which is inoculated by lactic acid bacteria. Lactic acid bacteria were isolated from Dolsan leaf mustard Kimchi (DLMK) at 20°C. In the optimum ripening period, the population ofLeuconostoc andLactobacilli in the DLMK were found to be high. TheLeuconostoc, Lactobacilli andLactococci strains were identified asLeuconostoc mesenteroides., Leuconostoc gelidum, Weissella confusa, Lactobacillus plantarum, Lactobacillus raffinolactis, Lactococcus lactis andWeissella confusa using the Biolog system. The most predominant strain which was isolated from DLMK wasWeissella confusa. As the results of the phylogenetic analysis using 16s rDNA sequence, theWeissella confusa turned out to beWeissella kimchii, with 99.0% similarity. To investigated the change of physiological activity in DLMJ by lactic acid bacteria, 7 predominant strains inoculated to DLMJ (Dolsan Leaf Mustard Juice). The cytotoxicity was found to be under 19.55% all cases Also, the antioxidative activity of the DLMJ treated with lactic acid bacteria was very low, which might have been due to the reduced antioxidative phytochemicals during the preparation of the sterile, sample. The ACE inhibiting activity of DLMJ by inoculation withWeissella kimchii was shown to be the highest (94.0%). This could be that the degradation of sulfur containing materials in DLMJ byWeissella kimchii gave rise to ACE inhibiting activity.     

Experiences with the use of a starter culture in the fermentation of maize for ‘kenkey’ production in Ghana

Controlled fermentation of maize was carried out using six strains of Lactobacillus fermentum and one strain of yeast, Saccharomyces cerevisiae, isolated from traditionally fermented maize dough as starter cultures for inoculum enrichement. The fermentations were monitored by pH, acidity, microbiological analysis and taste panel evaluation of two products, kenkey and koko, prepared from the fermented doughs. The strains of L. fermentum used as starter culture dominated the microflora during fermentation and in most inoculated doughs the required pH was attained by 24 h instead of 48 h of dough fermentation. Higher contents of lactic acid bacteria and yeasts were observed in inoculated doughs at the initial stages of fermentation but the spontaneously fermented doughs attained similar lactic acid bacteria and yeasts counts by 24 h of dough fermentation. The organoleptic quality of kenkey and koko prepared from doughs fermented with starter culture for 48 h was not significantly different from the traditional products. Kenkey prepared from doughs fermented for 24 h with starter culture were found to be unacceptable by the taste panel although similarly produced koko was acceptable.The authors are with the Food Research Institute, Council for Scientific and Industrial Research, P.O Box M 20. Accra, Ghana.     

Survey of bovine mycotic mastitis in dairy herds in the State of São Paulo,Brazil

The purpose of this study was to isolate fungi from the quarter milk of cow udders from several dairy herds and to identify the different genera and species involved in mastitis. A total of 2078 milk samples from normal, clinical and subclinical mastitis quarters from 22 dairy herds of 16 districts in the State of São Paulo, Brazil was utilized in this survey. Two hundred and fifty one (12.07%) fungi were isolated from the samples. Two hundred and eight of these (82.86%) were yeasts and 30 (11.95%) were moulds. The fungi were isolated in pure culture (24.77%) or in cultures mixed with bacteria (72.22%). The yeasts isolated were:Cryptococcus spp. (71 strains),Rhodotorula spp. (40),Candida spp. (68),Trichosporon cutaneum (21),Aureobasidium pullulans (7), andPichia ohmeri (1). Moulds classified in following genera were also isolated:Aspergillus (3),Penicillium (3),Alternaria (3),Phoma (3),Epicoccum (2), andGeotrichum (16).     

Enhanced production of lactic acid through the use of a novel aqueous two-phase system as an extractive fermentation system

 In order to enhance the productivity of lactic acid and reduce the end-product inhibition of fermentation, the partitioning and growth of four different strains of lactic acid bacteria in three different aqueous two-phase systems were studied. Polyethyleneglycol/ dextran, polyethyleneglycol/hydroxypropyl starch polymer (HPS), and a random copolymer of ethylene oxide and propylene oxide (EO-PO)/HPS were used as polymer systems. One strain each of Lactococcus lactis subsp. lactis and of Lactobacillus delbrueckii subsp. delbrueckii partitioned completely to the interface and bottom phase in two-phase systems with low polymer concentrations of EO-PO/HPS100 and EO-PO/ HPS200. The growth and production of lactic acid by two of three L. lactis strains in a two-phase system with 5.5% (w/w) EO-PO and 12.0% (w/w) HPS100 were reduced by less than 10% compared with a reference fermentation in a normal growth medium. The viability of L. lactis subsp. lactis ATCC 19435 was maintained for at least 50 h and with four top-phase replacements during extractive fermentation in the EO-PO/HPS100 system. Moreover, when cell density reached the stationary phase in the first extractive fermentation, the lactate production in this aqueous two-phase system was maintained. Received: 2 October 1995/Received revision: 16 January 1996/Accepted: 22 January 1996     

Characteristics of the bacteriocin produced by <Emphasis Type="Italic">Lactococcus lactis</Emphasis> subsp. <Emphasis Type="Italic">cremoris</Emphasis> CTC 204 and the effect of this compound on the mesophilic bacteria associated with raw beef

Summary >Screening for the bacteriocin production of strains of lactic acid bacteria from various meat and meat products resulted in the detection of a bacteriocin-producing Lactococcus lactis subsp. cremoris CTC 204, isolated from chicken. The bacteriocin inhibited not only closely related lactic acid bacteria (Lactobacillus helveticus), but also pathogenic microorganisms (Staphylococcus aureus, Listeria monocytogenes, Bacillus cereus, and Clostridium perfringens). It was inactivated by α-chymotrypsin, ficin, papain, and pronase E, but not by lipase or pepsin. This compound was heat stable even at autoclaving temperature (121°C for 10min) and was produced during refrigerated storage. It was also active over a wide pH range (2–10), but the highest activity was observed in the lower pH range. The results indicated that dipping raw beef in the bacteriocin produced by strain CTC 204 could contribute to the extension of the shelf life of refrigerated bovine meat.     

Use of starter culture of Lactobacillus plantarum BP04 in the preservation of dining-hall food waste

In this work, Lactobacillus plantarum BP04 was employed as starter culture in dining-hall food waste storage with different inoculant levels at 0, 2 and 10% (v/w) to suppress the outgrowth of pathogenic and spoilage bacteria. Inoculation by Lactobacillus plantarum BP04 was effective in accelerating pH drop and reducing the growth period of enterobacteria to 9, 7 and 2 days, corresponding to inoculant levels at 0, 2 and 10% (v/w). Increasing inoculum levels were found to inhibit the growth of Lactobacillus brevis and Leuconostoc lactis. HPLC analysis revealed that lactic acid was the predominant organic acid during the treatment of dining-hall food waste. Its concentration varied among the fermented processes reflecting variations of microbial activity in the fermented media.     

Utilization of cyanobacterial biomass from water bloom for bioproduction of lactic acid

Cyanobacterial biomass obtained from water blooms was successfully utilized as a material for lactic acid production. The starch contained in the biomass could be converted to D- and L-lactic acid with 80–90% yield by Lactobacillus amylovorus, in a manner similar to that contained in laboratory-cultured cyanobacterial biomass. The starch was also available for L-lactic acid production with similar high yields by L. agilis and L. ruminis that specifically produce L-lactic acid. The lactic acid production from the cyanobacterial biomass did not require any supplements such as yeast extract which are essential for lactic acid production from reagent soluble starch, indicating that nutrients contained in the cyanobacterial biomass might be effectively used for the production instead of the supplements. The starch content of the fresh cyanobacterial biomass from water bloom was increased from 10 to 19 and 24% by cultivation in 1 and 5% CO₂ in air, respectively. Using such starch-rich biomass, the concentration of lactic acid produced was successfully increased without changes in the conversion yield. These results indicate that wastewater bloom cyanobacteria could be utilized for the production of a useful compound, lactic acid.     

Effects of pressure on cell morphology and cell division of lactic acid bacteria

The effect of pressure and temperature on the growth of the mesophilic lactic acid bacteria Lactococcus lactis and Lactobacillus sanfranciscensis was studied. Both strains were piezosensitive. Lb. sanfranciscensis failed to grow at 50 MPa and the growth rate of Lc. lactis at 50 MPa was less than 30% of that at atmospheric pressure. An increase of growth temperature did not improve the piezotolerance of either organism. During growth under high-pressure conditions, the cell morphology was changed, and the cells were elongated as cell division was inhibited. At atmospheric pressure, temperatures above the optimal temperature for growth caused a similar effect on cell morphology and cell division in both bacteria as that observed under high-pressure conditions. The segregation and condensation of chromosomal DNA were observed by DAPI staining and occurred normally at high-pressure conditions independent of changes in cell morphology. Immunofluorescence microscopy of Lc. lactis cells demonstrated an inhibitory effect of high pressure on the formation of the FtsZ ring and this inhibition of the FtsZ ring formation is suggested to contribute to the altered cell morphology and growth inhibition induced by high pressure.Communicated by K. Horikoshi     

Vaginal lactic acid bacteria in the mare: evaluation of the probiotic potential of native Lactobacillus spp. and Enterococcus spp. strains

Lactic acid bacteria (LAB) are important members of the human vaginal microbiota and their presence is considered beneficial. However, little is known about native vaginal bacteria in other animal species such as the horse. The aim of this work was to quantify the vaginal lactic acid bacteria and lactobacilli of mares and to establish if selected equine vaginal lactic acid bacteria, particularly Lactobacillus and Enterococcus spp. strains, could exhibit potential as probiotics. The vaginal lactic acid bacteria and lactobacilli of 26 mares were quantified by plate counts. Five strains (three Lactobacillus spp. and two Enterococcus spp.) were characterised and adhesion to vaginal epithelial cells, antimicrobial activity and ability to form biofilms were evaluated. Lactic acid bacteria were recovered from the 26 samples and lactobacilli counts were detected in 18 out of 26 mares (69%). Probiotic properties tested in this study varied among the isolates and showed promising features for their use as equine probiotics.     

降解尿酸乳酸菌复合菌系的筛选与菌株组合提升降解效果

【背景】高尿酸症由血液中尿酸含量明显升高而导致,利用乳酸菌对人体的益生作用缓解高尿酸血症越来越受到关注。【目的】获得具有降解尿酸能力的乳酸菌复合菌系与纯培养菌株。【方法】以泡菜为样品来源,以尿酸为底物,采用MRS培养基筛选降解尿酸的乳酸菌复合菌系,通过高效液相色谱法测定复合菌系对尿酸的降解能力。【结果】得到一组乳酸菌复合菌系,当培养温度为37 °C、pH值为6.20、静置培养72 h后复合菌系对尿酸的降解率为12.08%;通过优化培养条件,当该菌系在以牛肉膏为单一氮源、初始pH值为5.00、温度为35 °C的条件下培养72 h,尿酸降解率上升至17.19%,降解率比优化前提高了42.3%;从该菌系中分离出两株具有尿酸降解能力的菌株UA-1与UA-2,它们的尿酸降解率分别为10.85%和8.65%;通过形态学观察和16S rRNA基因序列分析,经鉴定两株菌均为布氏乳杆菌(Lactobacillus buchneri)。将两株单菌组合降解尿酸试验发现,UA-1与UA-2比例为2:1的尿酸降解率为20.2%,比原复合菌系的降解能力提高了67.22%。【结论】研究证明了乳酸菌复合菌系对尿酸的降解能力优于单个菌株,为后续利用乳酸菌复合菌系应用提供了数据支持。     

Degradation of starchy substrates by a crude enzyme preparation and utilization of the hydrolysates for lactic fermentation

An enzyme preparation obtained from Aspergillus ustus, possessing cellulase, α-amylase, amyloglucosidase, proteinase and d-xylanase activities, was used along with commercial bacterial α-amylase and amyloglucosidase for the degradation of ragi (Eleusine coracana) flour and wheat (Triticum vulgare) bran. Lactic acid yield from ragi hydrolysate, adjusted to 5% reducing sugars (w/v), was 25% when fermented with Lactobacillus plantarum. The yields increased to 78% and 94% when the ragi hydrolysate was fortified with 20% and 60% (v/v) wheat bran hydrolysate, respectively. When commercial α-amylase and amyloglucosidase were used for the hydrolysis of ragi and wheat bran and L. plantarum was employed to ferment the hydrolysates containing 5% reducing sugars (w/v), lactic acid yields were 10% in ragi hydrolysate and 57% and 90% when the ragi hydrolysate was fortified with 20% and 60% (v/v) of wheat bran hydrolysate, respectively. α-Amylase and amyloglucosidase hydrolysed wheat bran added at 20% (v/v) as the sole source of nutrient to soluble starch hydrolysate (5% reducing sugars) gave 22% yield of lactic acid. The yield increased to 55% by the utilization of A. ustus enzyme preparation in addition to α-amylase and amyloglucosidase for wheat bran hydrolysis.     

Effect of Spirulina platensis biomass on the growth of lactic acid bacteria in milk

The stimulatory effect of aqueous suspensions of Spirulina platensis dry biomass extracted at pH 6.8 and 5.5 was studied on four lactic acid bacteria (LAB) grown in milk. The addition of dry S. platensis to milk (6 mg/ml) stimulated growth of Lactococcus lactis by 27%. The growth of other strains was also promoted. This revised version was published online in July 2006 with corrections to the Cover Date.