Characterization of bacteriocin-like inhibitory substances (BLIS) from lactic acid bacteria isolated from traditional Azerbaijani cheeses

The lactic acid bacteria (LAB) of southern Caucasus region present a special interest due to the diversity of lactic flora used for fermentations by various local populations during thousands of years. Four LAB strains, not identified previously, isolated from Motal and Brunza typical Azerbaijani cheeses were subjected to phenotypic identification and three of them could be identified as Lactobacillus paracasei subsp. paracasei and one as Lactobacillus rhamnosus. Test strains such as Lactobacillus bulgaricus 340 and Saccharomyces cerevisiae were inhibited by the four isolated strains. Antibacterial activity against Escherichia coli HB 101 was detected in Lactobacillus paracasei BN ATS 8w and L. rhamnosus FAZ 16m. L. paracasei BN ATS 5w and 8w, and L. rhamnosus FAZ 16m showed inhibitory effect on Staphylococcus aureus Cip 9973. The inhibition of Candida pseudotropicalis was detected only when using L. paracasei species BN ATS 5w and 7w. Culture of Listeria innocua was insensitive to the antimicrobial substances of all the studied strains. Complete inactivation or significant reduction in antibacterial activity was observed after treatment of cell-free supernatants with pronase E and proteinase K, but not with trypsin (except for L. rhamnosus FAZ 16m), indicating the protein nature of the active agents. Amylase treatment totally inactivated the substances of L. paracasei, what implies the importance of glycosylation for the activity. The activities of all the bacteriocin-like substances from studied LABs were stable over a wide pH range from 3 to 11.     

Culture-independent detection of microorganisms in traditional Slovakian bryndza cheese

In order to investigate the microflora of Slovakian bryndza cheese (a cheese containing unpasteurized or pasteurized ewes' milk component) by a culture-independent method, DNA was extracted directly from 7 bryndza samples and analysed by an innovative method. Using the universal prokaryotic and fungal primers, ribosomal DNA internal transcribed spacer (ITS) regions with variable length were amplified. The standard universal reverse primer L1 aligning to bacterial 23 s rDNA was found unsuitable for some lactic acid bacteria and other species based on in silico analysis. Therefore, L1 primer was replaced by a combination of novel primers GplusR and GminusR aligning to the adjacent, more conserved DNA region. The amplification profiles were visualised by both standard electrophoresis and by fluorescent capillary gel electrophoresis. From representative samples, major amplicons were excised from the gel, cloned and sequenced. Sequencing revealed that the samples contained Lactobacillus delbrueckii, Lactobacillus brevis, Streptococcus thermophilus, Lactococcus lactis, Lactococcus raffinolactis, Streptococcus macedonicus, Leuconostoc pseudomesenteroides, Debaromyces hansenii, Mucor fragilis, Yarrowia lipolytica and Galactomyces geotrichum. These results represent an extension of the knowledge on the microflora of Slovakian bryndza cheese. The introduced automated ribosomal DNA intergenic spacer analysis of the bacterial and fungal genomes proved to be very effective in the application of studying microflora of cheese.     

豆乳的乳酸菌发酵初探

利用乳酸菌发酵豆乳,结果表明,如同牛乳一样,豆乳也能提供乳酸菌生长所需的营养物质,并使豆乳的PH下降和产生凝乳现象。但经乳酸菌发酵的豆乳,其酸度远不如酸乳,而且乳酸菌的生长量也低于牛乳(前者仅10~7个/ml,后者达10~8个/ml以上)。添加葡萄糖、乳糖等物质于豆乳中,虽有助于提高豆乳的酸度,但效果不甚显著。此外,豆乳经乳酸菌发酵后也不能产生酸牛奶所具有的特殊香味或风味,也未检出双乙酰(丁二酮)和乙醛等物质。     

Proteolysis in model Portuguese cheeses: Effects of rennet and starter culture

To shed further light onto the mechanisms of proteolysis that prevail throughout ripening of Portuguese cheeses, model cheeses were manufactured from bovine milk, following as much as possible traditional manufacture practices – using either animal or plant rennet. The individual role upon proteolysis of two (wild) strains of lactic acid bacteria – viz. Lactococcus lactis and Lactobacillus brevis, which are normally found to high viable numbers in said cheeses, was also considered, either as single or mixed cultures. Our experimental results confirmed the influence of rennet on the proteolysis extent, but not on proteolysis depth. On the other hand, the aforementioned strains clearly improved release of medium- and small-sized peptides, and contributed as well to the free amino acid pool in cheese.     

Volatile sulphur compounds-forming abilities of lactic acid bacteria: C-S lyase activities

Volatile sulphur compounds (VSCs) are of prime importance in the overall aroma of cheese and make a significant contribution to their typical flavours. Thus, the control of VSCs formation offers considerable potential for industrial applications. Here, lactic acid bacteria (LAB) from different ecological origins were screened for their abilities to produce VSCs from L-methionine. From the data presented, VSC-forming abilities were shown to be strain-specific and were correlated with the C-S lyase enzymatic activities determined using different approaches. High VSCs formation were detected for those strains that were also shown to possess high thiol-producing abilities (determined either by agar plate or spectrophotometry assays). Moreover, differences in C-S lyase activities were shown to correspond with the enzymatic potential of the strains as determined by in situ gel visualization. Therefore, the assessment of the C-S lyase enzymatic potential, by means of either of these techniques, could be used as a valuable approach for the selection of LAB strains with high VSC-producing abilities thus, representing an effective way to enhance cheese sulphur aroma compounds synthesis. In this regard, this study highlights the flavour forming potential of the Streptococcus thermophilus STY-31, that therefore could be used as a starter culture in cheese manufacture. Furthermore, although C-S lyases are involved in both biosynthetic and catabolic pathways, an association between methionine and cysteine auxotrophy of the selected strains and their VSCs-producing abilities could not be found.     

多菌种混合乳酸菌发酵酸豆奶工艺研制

本文研究了市售酸牛奶和自制酸泡菜中分离筛选菌种,确定菌种间的比例。通过对生产过程中的条件进行初步摸索,以鲜豆奶为主要原料,采用多菌种混合液态发酵,可生产出色、香、味俱佳,不舍任何防腐剂的纯天然酸豆奶新产品,从而为工业化生产乳酸菌发酵酸豆奶提供技术依据。     

发酵型营养米乳的研制

以大米为原料,经过糊化等预处理后接入一定的乳酸菌进行发酵,最终获得了感官、组织状态良好的发酵型米乳。本研究对大米的加水量、菌种的种类及比例、发酵条件等进行了选择和优化,确定了最佳工艺条件。     

微生物抑制法快速检测鲜牛乳中红霉素残留

目的利用微生物抑制法快速检测鲜牛乳中红霉素残留。方法将待测的牛奶样品和空白样品经杀菌后接入乳酸菌,经过相同时间发酵后,计算待测牛奶样品发酵前后的pH变化差值与空白样品发酵前后的pH变化差值,从而判断鲜牛奶中红霉素残留的含量。结果经过单因素及正交实验,确定乳酸菌发酵产酸条件如下:接种量0.15 U/L、发酵温度40℃、菌粉溶解时间25 min、发酵时间3.5 h,利用乳酸菌发酵产酸建立了检测鲜牛奶中红霉素残留的标准曲线Y=0.024X-0.013(其中,Y为△pH,X为鲜牛乳样品中的红霉素浓度),线性相关系数R~2=0.9949,检测的灵敏度IC_(50)为29.71μg/L,检测的线性范围为0~50μg/L,加标回收率为103.54%~124.58%,同一批次内检测的变异系数为4.39%~16.46%。结论建立的检测方法可用于筛选鲜牛乳中是否有抗生素残留并进行定量检测。     

乳酸发酵豆乳饮料生产工艺

以大豆为主要原料,经磨浆、加入驯化后的乳酸菌发酵成熟后进行调配,制成乳酸发酵豆乳饮料。     

Evaluation of genetic polymorphism among Lactobacillus rhamnosus non-starter Parmigiano Reggiano cheese strains

Parmigiano Reggiano (PR) is an Italian cooked, long-ripened cheese made with unheated cow's milk and natural whey starter. The microflora is involved in the manufacturing of this cheese, arising from the natural whey starter, the raw milk and the environment. Molecular studies have shown that mesophilic non-starter lactic acid bacteria (NSLAB) are the dominant microflora present during the ripening of PR. In this study, a characterisation of Lactobacillus rhamnosus isolated from a single PR manufacturing and ripening process is reported, using a combination of genotypic fingerprinting techniques (RAPD-PCR and REP-PCR). The intraspecies heterogeneity evidenced for 66 strains is correlated to their abilities to adapt to specific environmental and technological conditions. The detection of biotypes that correlate with specific moments in cheese ripening or differential development throughout this process suggests that these strains may have specific roles closely linked to their peculiar technological properties.     

Diversity of lactic acid bacteria population in ripened Parmigiano Reggiano cheese

The diversity of dominant lactic acid bacteria population in 12 months ripened Parmigiano Reggiano cheeses was investigated by a polyphasic approach including culture-dependent and independent methods. Traditional plating, isolation of LAB and identification by 16S rDNA analysis showed that strains belonging to Lactobacillus casei group were the most frequently isolated. Lactobacillus helveticus, Lactobacillus delbrueckii subsp. lactis, Lactobacillus parabuchneri, and Lactobacillus buchneri species were detected with lower frequency. PCR-denaturing gradient gel electrophoresis (DGGE) applied to DNA extracted directly from cheese samples and sequencing of rDNA amplicons confirmed the complex microbiological pattern of LAB in ripened Parmigiano Reggiano cheeses, with the significant exception of the Lactobacillus fermentum species, which dominated in several samples, but was not detected by cultivation. The present combination of different approaches can effectively describe the lactic acid bacteria population of Parmigiano Reggiano cheese in advanced stages of ripening, giving useful information for elucidating the role of LAB in determining the final cheese quality.