Lactic Acid Bacteria Cultures Research Articles

In vitro Studies on Antimicrobial Activities of Lactic Acid Bacteria Isolated from Fresh Vegetables for Biocontrol of Tomato Pathogens

Aims: This study was focused on using Lactic Acid Bacteria (LAB) isolated from fresh vegetables which has been molecularly identified for in vitro control of some tomato pathogens. Study Design: The inhibitory potentials of supernatant obtained from previously characterized LAB isolates or vegetable origin were investigated against some tomato phytopathogens using agar-well method with the view to develop biological agents for some tomato disease causing organisms. Place and Duration of Study: Biotechnology Centre of Federal University of Agriculture, Abeokuta, Ogun State, Nigeria, between January 2011 and February 2012. Methodology: The antimicrobial activities of LAB against some tomato phytopathogenic bacteria which include (Xanthomonas campestries, Erwinia caratovora, and Pseudomonas syringae) were obtained by using the agar well diffusion method. Results: The result indicates that cell free culture of LAB from fresh vegetables origin Original Research Article British Microbiology Research Journal, 4(3): 351-359, 2014 352 (Weissella paramesenteroides, Lactobacillus pentosus, Weissella cibaria, Pediococcus pentosaceus, Weissella kimchi and Lactobacillus plantarum) can inhibits these bacteria by creating clear zones of inhibition around the wells containing cell free supernatants of the above mentioned strains of lactic acid bacteria. Pediococcus pentosaceus showed the highest zone of inhibition against Xanthomonas campestries at 15 mm radius, Weissella kimchi was the least effective against Pseudomonas syringae at 3.67 mm and Erwinia caratovora at 3.50 mm radius. Conclusion: Tomato disease causing organisms can be most likely biologically controlled by using extracts from LAB. This finding will reduce the potential hazard from the use of chemical herbicides on plant.

Modulation of gut microbiota associated with abalone Haliotis gigantea by dietary administration of host-derived Pediococcus sp. Ab1

In the present study, we evaluated how dietary administration of host-derived Pediococcus sp. Ab1 has an effect on the abalone gut microbiota using a culture-dependent method and 16S rRNA gene library analysis. The culturable lactic acid bacteria number in the probiotic sample was 105 higher than that in the non-probiotic sample, and we speculate that this significant increase was due to colonization of Ab1 into abalone gut. The result of a culture-dependent method showed that the proportion of Vibrio halioticoli clade, which is known to be a beneficial resident bacterium to abalone, was much higher in the probiotic sample than in the non-probiotic sample. 16S rRNA gene clone sequences revealed that gut microbiota in the probiotic sample was obviously diverse compared to the non-probiotic sample, probably due to improvement of the gut environment by Ab1 colonization. In addition, some beneficial bacteria–like sequences such as V. halioticoli were only found in the probiotic sample. These results suggest that the dietary administration of Ab1 to abalone gut has a great effect on modulation of not only culturable but also unculturable gut microbiota. Our results are useful for future investigations into understanding the effect of probiotics on gut microbiota.

Cheddar Cheese Ripening and Flavor Characterization: A Review

Cheddar cheese is a biochemically dynamic product that undergoes significant changes during ripening. Freshly made curds of various cheese varieties have bland and largely similar flavors and aroma and, during ripening, flavoring compounds are produced that are characteristic of each variety. The biochemical changes occurring during ripening are grouped into primary events including glycolysis, lipolysis, and proteolysis followed by secondary biochemical changes such as metabolism of fatty acids and amino acids which are important for the production of secondary metabolites, including a number of compounds necessary for flavor development. A key feature of cheese manufacture is the metabolism of lactose to lactate by selected cultures of lactic acid bacteria. The rate and extent of acidification influence the initial texture of the curd by controlling the rate of demineralization. The degree of lipolysis in cheese depends on the variety of cheese and may vary from slight to extensive; however, proteolysis is the most complex of the primary events during cheese ripening, especially in Cheddar-type cheese.

A comprehensive investigation into sample extraction and method validation for the identification of antifungal compounds produced by lactic acid bacteria using HPLC-UV/DAD

High Performance Liquid Chromatography with Ultra Violet/Diode Array Detection (HPLC-UV/DAD) is an analytical tool that has the potential to develop and advance analytical research and routine analyses in industrially-important Lactic Acid Bacteria (LAB) with the aim of detecting metabolites with antifungal capabilities. Diode array detection is a multi-wavelength detection system that enables the simultaneous collection of absorption data from a range of UV and visible (UV/vis) wavelengths. The majority of antifungal organic compounds have characteristic UV/vis absorption spectra due to the presence of chromophoric groups or structures, this feature can be utilised to assist in identifying and quantifying the components (known and unknown) as they pass through the detector. HPLC-UV/DAD is robust, cheap to purchase and run, easy to maintain, the software is user friendly and automated for quantitation and the methodology can be easily switched/up-scaled to semi-preparative applications for target analyte isolation and purification. However inefficient sample preparation has become the limiting factor for the routine use of HPLC-UV/DAD in the LAB area. This manuscript will outline the development and validation of an HPLC-UV/DAD method that addresses all of the factors that has limited the routine use of this technology for LAB samples. The method has been developed to quantitatively profile fifteen antifungal compounds in LAB strains in a run time of 40 minutes, allowing the assessment of crude samples (direct injection), eliminating sample matrix chromatographic baseline rise effects through the optimisation of liquid–liquid extraction (LLE with average recoveries = 87%) and assessing a widely applied SPE methodology that is shown to yield poor recoveries for the suite of target analytes. The method reported is validated (linear calibration curve: 1–50 μg mL−1; R2 ≥ 0.999; intra-day repeatability gave RSD values (n = 3) ranging from 0–8.8%; inter-day repeatability gave RSD values (n = 9) ranging from 0.3–7.9%; LODs of 0.2–0.6 μg mL−1 and LOQs of 0.5–0.9 μg mL−1 following LLE applied to a range of crude culture broths and extracted LAB cultures, and all results were confirmed against a liquid chromatography (LC) linear ion trap quadrupole (LTQ) Orbitrap hybrid Fourier transform mass spectrometer (FTMS). In summary, through this methodology, HPLC-UV/DAD can be routinely used as a quantitative screening technique for both culture broth (e.g. de Man, Rosaga and Sharpe (MRS)) and LLE-extracted LAB cultures.

Phenotypic and genotypic characterization of some lactic Acid bacteria isolated from bee pollen: a preliminary study.

In the present work, five hundred and sixty-seven isolates of lactic acid bacteria were recovered from raw bee pollen grains. All isolates were screened for their antagonistic activity against both Gram-positive and Gram-negative pathogenic bacteria. Neutralized supernatants of 54 lactic acid bacteria (LAB) cultures from 216 active isolates inhibited the growth of indicator bacteria. They were phenotypically characterized, based on the fermentation of 39 carbohydrates. Using the simple matching coefficient and unweighted pair group algorithm with arithmetic averages (UPGMA), seven clusters with other two members were defined at the 79% similarity level. The following species were characterized: Lactobacillus plantarum, Lactobacillus fermentum, Lactococcus lactis, Pediococcus acidilactici, Pediococcus pentosaceus, and unidentified lactobacilli. Phenotypic characteristics of major and minor clusters were also identified. Partial sequencing of the 16S rRNA gene of representative isolates from each cluster was performed, and ten strains were assigned to seven species: Lactobacillus plantarum, Lactobacillus fermentum, Lactococcus lactis, Lactobacillus ingluviei, Pediococcus pentosaceus, Lactobacillus acidipiscis and Weissella cibaria. The molecular method used failed to determine the exact taxonomic status of BH0900 and AH3133.

Toward Probiotict Food Product from Meat Through Isolation and Identification Lactic Acid Bacteria As Probiotic Culture Stater

Probiotic food products of meat can provide extensive benefits, to increase the shelf life and nutritional value also improve the taste. The use of lactic acid bacteria culture (LAB) derived from the isolation of the meat and the addition of probiotic cultures (Lactobacilli and Bifidobacteria) in fermented sausage processing is expected to produce a probiotic sausage products with nutritional value, and better shelf life and improve health. This study aimed to isolate and identify lactic acid bacteria (LAB) of meat as a starter culture fermented sausages. The parameters observed were gram test, catalase, motility, gas production, type of fermentation, growth at various temperatures and pH. The results were obtained 28 isolates, 17 isolates were able to produce acid and 8 of them are lactic acid bacteria (LAB) with the characteristics of gram-positive, catalase negative, not motile, produces gas, are hetero and homo fermentative, optimum growth temperature of 300C and a few of them are able to grow on pH 3.5. Lactic acid bacteria that able to be combined with probiotics as sausage starter culture to the probiotic food products of meat.

Diversity of Lactic Acid Bacteria in the Korean Traditional Fermented Beverage Shindari, Determined Using a Culture-dependent Method

The fermented food Shindari is a low-alcohol drink that is indigenous to Jeju island, South Korea. In this study,the diversity of lactic acid bacteria (LAB) in Shindari was determined using a culture-dependent method. LAB were culti-vated from Shindari samples using two different LAB culture media. Twenty-seven strains were randomly selected and iden-tified by 16S rRNA gene sequence analysis. The identified LAB strains comprised 6 species within the Enterococcus,Lactobacillus and Pediococcus genera. Five of the species, namely Enterococcus faecium, Lactobacillus fermentum, L. plan-tarum, Pediococcus pentosaceus and P. acidilactici were isolated from MRS medium, while 1 species, L. pentosus, was iso-lated from Rogosa medium. Most of the isolated strains were identified as members of the genus Lactobacillus (78%). Thisstudy provides basic microbiological information on the diversity of LAB and provides insight into the ecological roles ofLAB in Shindari.

Increased viability of probioticLactobacillus rhamnosusafter osmotic stress

The objective of this work was to assess the impact of various NaCl concentrations on Lactobacillus rhamnosus OXY viability after freeze-drying. Osmotic stress was applied during the exponential growth phase of bacterial culture. At salt concentrations between 0.2–0.5 M, a high biomass concentration and a significant increase in cell viability after lyophilisation was observed. An analysis of two-dimensional protein gels indicated the presence of shock proteins, for example, GroEL, ClpB, DnaK, TF, which provide resistance during freeze-drying and subsequent storage. On the basis of these results, it is recommended that lactic acid bacteria cultures be sub-lethally treated with 0.5 M NaCl before freeze-drying.

Molecular characterisation and antimicrobial activity of bacteria associated with submerged lactic acid cassava fermentation

Molecular identification of microorganisms associated with submerged cassava fermentation was carried out and isolates of lactic acid bacteria (LAB) were examined for antimicrobial activity, including ability to produce antimicrobial peptides as a first step to define starter cultures for controlled cassava fermentations. A total of 75 isolates, including 41 LAB, 31 aerobic bacteria (AB) and three anaerobic bacteria were isolated from unfermented and fermenting cassava roots, cassava leaves and fermented cassava dough and identified by a combination of phenotypic tests and sequencing of 16S rRNA, rpoA, rpoB and pheS genes. Microbial diversity at interspecies and intraspecies level was screened by, respectively, PCR amplification of the 16S-23S rDNA intergenic transcribed spacer (ITS-PCR) and repetitive sequence based PCR (rep-PCR). Antimicrobial activity of LAB cultures and supernatants against indicator bacteria; Escherichia coli, Salmonella enterica serotype Typhimurium (S. Typhimurium), Bacillus cereus and Staphylococcus aureus was studied using agar diffusion tests. Furthermore, inactivation of indicator bacteria was investigated in both liquid medium and during controlled cassava fermentation. Results revealed a diversity of bacterial genera, species and subspecies associated with submerged cassava fermentation. DNA sequencing enabled identification of LAB isolates as Lactobacillus plantarum, Weissella confusa, Weissella paramesenteroides, Lactobacillus rhamnosus, Lactobacillus hilgardii, Lactobacillus paracasei, Leuconostoc mesenteroides, Enterococcus faecium, Enterococcus casseliflavus, and Pediococcus acidilactici. Lactobacillus spp. were the predominant LAB and were present in all cassava samples studied. Aerobic bacteria were predominantly Bacillus spp., including Bacillus subtilis, Bacillus amyloliquefaciens and B. cereus. Other species identified included Staphylococcus pasteuri and Clostridium beijerinkii. Cells, supernatants and cell free supernatants (CFS) of selected LAB isolates were able to inhibit both Gram positive and Gram negative pathogenic bacteria. LAB isolates inactivated all indicator organisms during controlled cassava fermentations, with a 4–6 log reduction after 48 h fermentation. The antimicrobial effect of the LAB was attributed to acid production.

Isolation, Identification and Characterization of Lactic Acid Bacteria From Shao-jiou-luo (Fermented Zoned Cerith), a Traditional Fermented Food in Taiwan

This study investigated the distribution of the native lactic acid bacteria (LAB) population in shao-jiou-luo (fermented zoned cerith), a popular traditional fermented food in Taiwan. A total of 129 cultures of LAB were isolated from shao-jiou-luo samples, and 14 cultures were isolated from its main raw material, zoned cerith. These isolates were divided into classes by phenotype and then into groups by restriction fragment length polymorphism analysis and sequencing of 16S ribosomal DNA. The identification results revealed 11 bacterial species belonging to five genera in shao-jiou-luo and three species belonging to two genera in zoned cerith. The most common bacterial genera in shao-jiou-luo were heterofermentative Weissella and Leuconostoc. On the other hand, Enterococcus aquimarinus was the most common species found in the raw zoned cerith sample. In conclusion, the results suggest that heterofermentative LAB species greatly contributed to the fermentation of shao-jiou-luo. This is the first report describing the distribution and varieties of LAB that exist in the shao-jiou-luo fermentation process and raw zoned cerith.

Diversity of Lactic Acid Bacteria during Fermentation of a Traditional Chinese Fish Product, Chouguiyu (Stinky Mandarinfish)

Chouguiyu, or stinky mandarinfish, is a traditional Chinese fermented fish product made of mandarinfish by spontaneous fermentation at the anaerobic condition with low-salt concentration. In order to get a primary understanding of the microbial community presenting in the Chouguiyu fermentation, 61 cultures of lactic acid bacteria (LAB) from various fermentation period were isolated using MRS agar plates and characterized based on a combination of phenotypic and genotypic approaches including amplified ribosomal DNA restriction analysis (ARDRA) and 16S rRNA partial gene sequencing analysis. Eight distinct bacterial species belonging to 6 genera were identified in total. Among them, Lactobacillus sakei was the dominant species (63%) during the fermentation, which exhibited great variety in phenotypic tests but unique genotypic characters. Meanwhile, the other LAB species including Lactococcus (Lc.) garvieae, Lc. lactis, Lc. raffinolactis, Vagococcus sp., Enterococcus hermanniensis, Macrococcus caseolyticus as well as Streptococcus parauberis were also recovered from the different fermentation periods, especially at the initial point of the fermentation. This seems to be the 1st report investigating the LAB composition involved in Chouguiyu fermentation and the data obtained in this study may be valuable for selecting starter culture for Chouguiyu industrial-scale production.

Population dynamics of mixed cultures of yeast and lactic acid bacteria in cider conditions

The objective of this work was to study the malolactic bioconversion in low acidity cider, according Brazilian conditions. The apple must was inoculated with Saccharomyces cerevisiae or S. cerevisiae with Oenococcus oeni. The control contained the indigenous microorganisms. Fermentation assays were carried out with clarified apple must from the Gala variety. At the beginning of fermentation, there was a fast growth of the non-Saccharomyces yeast population. Competitive inhibition occurred in all the assays, either with inoculated or indigenous populations of the yeast. The lactic acid bacteria count was ca. 1.41·102CFU/mL at the beginning and 106CFU/mL after yeast cells autolysis. The lactic bacteria O. oeni reached the highest population (107CFU/mL) when added to the apple must after the decline of the yeast. The malic acid was totally consumed during the alcoholic fermentation period (80.0 to 95.5 %) and lactic acid was still synthesized during the 35 days of malolactic fermentation. These results could be important in order to achieve a high quality brut, or sec cider obtained from the dessert apple must.

Isolation and identification of lactid acid bacteria originated from king grass (Pennisetum purpureophoides) as candidate of probiotic for livestock

A study was conducted to isolate and identify strain of lactic acid bacteria (LAB) isolated from king grass, and to determine their potential as candidate of probiotic for livestock. The LAB was isolated by culturing king grass extract in De Man, Rogosa and Sharpe (MRS) medium. The pure culture LAB was used to identify strain of bacteria using Analytical Profile Index (API) 50 CH kit. The result showed that the strain bacteria was identified as Lactobacillus plantarum. L. plantarum was able to survive in extreme condition at pH 2 and 0.3% bile salt. L . plantarum also survived against pathogenic bacteria i.e. Staphylococcus aureus , Escherechia coli and Salmonella thypi . It is concluded that L. plantarum isolated from king grass could potentially to be used as probiotic for livestock. Key Words : Bile salt, Lactic acid bacteria, Pathogen, Probiotic, pH.

폐배추 추출물을 이용한 Leuconostoc citreum GR1 종균 배양용 최적 배지 및 배양 조건 개발

In the kimchi manufacturing process, the starter is cultured on a large-scale and needs to be supplied at a low price to kimchi factories. However, current high costs associated with the culture of lactic acid bacteria for the starter, have led to rising kimchi prices. To solve this problem, the development of a new medium for culturing lactic acid bacteria was studied. The base materials of a this novel medium consisted of Chinese cabbage extract, a carbon source, a nitrogen source, and inorganic salts. The optimal composition of this medium was determined to be 30% Chinese cabbage extract, 2% maltose, 0.25% yeast extract, and 2× salt stock (2% sodium acetate trihydrate, 0.8% disodium hydrogen phosphate, 0.8% sodium citrate, 0.8% ammonium sulfate, 0.04% magnesium sulfate, 0.02% manganese sulfate). The newly developed medium was named MFL (medium for lactic acid bacteria). After culture for 24 hr at 30°C, the CFU/mL of Leuconostoc (Leuc.) citreum GR1 in MRS and MFL was 3.41×10 9 and 7.49×10 9 , respectively. The number of cells in the MFL medium was 2.2 times higher than their number in the MRS media. In a scale-up process using this optimized medium, the fermentation conditions for Leuc. citreum GR1 were tested in a 2 L working volume using a 5 L jar fermentor at 30°C. At an impeller speed of 50 rpm (without pH control), the viable cell count was 8.60×10 9 CFU/mL. From studies on pH-stat control fermentation, the optimal pH and regulating agent was determined to be 6.8 and NaOH, respectively. At an impeller speed of 50 rpm with pH control, the viable cell count was 11.42×10 9 (1.14×10 10 ) CFU/mL after cultivation for 20 hr - a value was 3.34 times higher than that obtained using the MRS media in biomass production. This MFL media is expected to have economic advantages for the cultivation of Leuc. citreum GR1 as a starter for kimchi production.

Physico‐Chemical Properties of Sourdough Bread Production Using Selected Lactobacilli Starter Cultures

AbstractThe acidification properties and technological performance (dough elasticity and bread volume) of four individual lactic acid bacteria (LAB) starter cultures (Lactobacillus delbrueckii subsp. bulgaricus LB18; Lactobacillus delbrueckii subsp. bulgaricus CNRZ 737; Lactobacillus delbrueckii subsp. bulgaricus 2483; Lactobacillus helveticus LH30) were investigated in relation to dough and bread quality derived from a sourdough fermentation. A plain dough mixture (without LAB starter cultures) was used as a control against the sourdough systems with added cultures. The exopolysaccharide (EPS) producing LAB starter cultures significantly increased dough stickiness and extendibility compared with the plain dough system. Total EPS production by the LAB cultures during sourdough formation differed depending on the culture used (in the order of NCFB 2483 > Z737 > LB18 > LH30). The viscoelastic nature of the dough was increased with the use of LAB cultures. Bread volume significantly increased with the inclusion of EPS cultures. Bread staling (as determined by the extent of firming of bread) was reduced with the use of ropy cultures. Increased exopolysaccharide production during dough formation appeared to be associated with reduced staling and increased bread volume.Practical ApplicationsSourdough bread and cereal products have experienced increased attention from consumers for either health or sensorial reasons. The current research investigate the potential use of a selected range of exopolysaccharide producing starter strains in an attempt to characterise textural and sensorial qualities derived from ropy and nonropy lactic acid bacteria (LAB) starter cultures. The results indicate a clear potential for these LAB cultures in improving consumer quality and shelf‐life stability of sourdough products.

Fermentation of non-sterilized fish biomass with a mixed culture of film-forming yeasts and lactobacilli and its effect on innate and adaptive immunity in mice

Non-sterilized fish waste containing fish bones was fermented using combined starter cultures of film-forming yeast (Candida ethanolica) and lactic acid bacteria (LAB; Lactobacillus casei and Lactobacillus rhamnosus) in order to obtain a liquefied fermented broth without spoiling. During the entire fermentation, the number of LAB cells was maintained at a high level (6 × 10(8)-5 × 10(7) cells/ml). Although the number of general bacteria was 10(6)cell/ml after adding non-sterilized fish biomass, its growth was suppressed to be 1-3 × 10(4) cells/ml. The entire biomass had completely liquefied and the fermented broth contained all 20 α-amino acids composed of protein and also various kinds of minerals in abundance. The weight of mice group fed the fermented broth content feed (sample feed) for 31 days significantly increased compared with that fed no broth feed (control feed) (21.37 g vs 20.76 g (p < 0.05). No abnormal behavior and appearance were observed. All internal organs (the heart, the liver, the lung, the intestines, and the spleen) of both groups were confirmed to be normal by visual observation. In peripheral blood, the percentages of NK cells and CD8+ T cells of the mice in the sample feed group increased significantly relative to those in the control feed group (NK cells: 19% vs 11%, CD8+ T cells: 9% vs 5%, p < 0.05). In the spleen, the percentage of NK cells in the sample feed group also increased significantly compared to that in the control feed group (p < 0.05). The fermented fish biomass is expected to be effective for innate and adaptive immunity and thus fit for animal feed.

Mixed cultures of Kimchi lactic acid bacteria show increased cell density and lactate productivity

This study was carried out to determine the characteristics of cell growth, lactate production and amino acid secretion among four kimchi lactic acid bacteria ( Leuconostoc mesenteroides JEI, Leuconostoc kimchi 132, Lactobacillus sakei 171, and Weissella koreensis 521) alone and in selected mixtures. In solo culture, L. sakei 171 was superior in cell growth, lactate production and the release of amino acids to the extracellular medium. In contrast, W. koreensis 521 showed the least cell growth, lactate production and amino acid release among the tested bacteria. W. koreensis 521 consumed essential amino acids for growth, whereas L. sakei 171 released several of the essential amino acids important for the growth of W. koreensis 521. When we mixed L. sakei 171 and W. koreensis 521 at optimal concentrations, the obtained cell growth and lactic acid production were higher than those seen with either strain alone, presumably reflecting mutual effects between the two strains. Mixed culture of two kimchi lactobacilli on batch fermentation increased the cell density and lactic acid production with low nutrients consumption. These results suggest that mixed culturing of kimchi lactobacilli may be more effective than single culturing of kimchi lactic acid bacteria for improving lactic acid production. Keywords : Kimchi lactic acid bacteria, amino acid utilization, nutrients consumption. African Journal of Biotechnology Vol. 12(25), pp. 4000-4005

Suitability of selected autochthonous lactic acid bacteria cultures for Pecorino Sardo Dolce cheese manufacturing: influence on microbial composition, nutritional value and sensory attributes

The use of an autochthonous starter culture made up by Streptococcus thermophilus, Lactococcus lactis subsp. lactis, Lactobacillus plantarum and Lactobacillus casei in the production of experimental Pecorino Sardo Dolce PDO resulted in a high presence of presumptive mesophilic lactococci and lactobacilli, ~10 log colony‐forming units (cfu), and in significantly higher amount of total free amino acids and free fatty acids, 195.1 vs 178.9 and 328.6 vs 191.8 mg/100 g of experimental vs control cheese (made using a commercial starter) respectively. The experimental cheese also displayed better sensory properties such as taste and aftertaste.

ISOLATION AND IDENTIFICATION OF MICROORGANISMS DURING SPONTANEOUS FERMENTATION OF MAIZE [Isolasi dan Identifikasi Mikroorganisme pada Fermentasi Spontan Jagung

Maize was traditionally the second most common staple food in Indonesia. Conversion to maize flour has been accomplished to improve its convenience. Traditionally, maize flour is produced by soaking the kernels in water followed by grinding. It was reported that final physicochemical characteristics of the maize flour were influenced by spontaneous fermentation which occurred during soaking. This research aimed to isolate and identify important microorganisms that grew during fermentation thus a standardized starter culture can be developed for a more controlled fermentation process. Soaking of maize grits was conducted in sterile water (grits:water=1:2, w/v) in a closed container at room temperature (±28ºC) for 72 hours. After 0, 4, 12, 24, 36, 48, 72 hours, water and maize grits were sampled and tested for the presence of mold, yeast, and lactic acid bacteria (LAB). Isolates obtained from the spontaneous fermentation were reinoculated into the appropriate media containing starch to observe their amylolytic activity. Individual isolate was then identified; mold by slide culture method, while yeast and LAB by biochemical rapid kits, i.e. API 20C AUX and API CH50, respectively. The number of each microorganism was plotted against time to obtain the growth curve of the microorganisms during spontaneous fermentation. The microorganisms were identified as Penicillium chrysogenum, P. citrinum, A. flavus, A. niger, Rhizopus stolonifer, R.oryzae, Fusarium oxysporum, Acremonium strictum, Candida famata, Kodamaea ohmeri, Candida krusei/incospicua, Lactobacillus plantarum 1a, Pediococcus pentosaceus, L. brevis 1, L. plantarum 1b, and L. paracasei ssp paracasei 3. Four molds and one yeast were amylolytic while none of the LAB was capable of starch hydrolysis. The growth curve suggested that the amylolitic mold and yeast grew to hydrolyze starch during the course of fermentation, while the LABs benefited from the hydrolyzed products and dominated the later stage of the fermentation.

A NMR metabolomics study of the ripening process of the Fiore Sardo cheese produced with autochthonous adjunct cultures

Fiore Sardo (FS) is a traditional Italian raw ewe’s milk cheese carrying a Protected Designation of Origin (PDO). This study investigated the kinetics of FS cheese ripening by physicochemical parameters, microbial counting, and NMR metabolomics using aqueous extracts. Four Fiore Sardo cheeses, manufactured from milk with deliberately added autochthonous lactic acid bacteria (LAB) or commercial starters were studied during a period of 90days of ripening. Major differences in the metabolic profiles were observed amongst the samples as a function of the adjunct culture utilised. 1H NMR metabolomics in combination with multivariate data analysis was able to classify cheese samples on the basis of their maturation age and the type of added cultures. These findings lay the metabolic basis for the authentication of Fiore Sardo cheese produced in compliance with PDO specifications which allow the use of only native LAB cultures.