Sensitivity of Molds From Spoiled Dairy Products Towards Bioprotective Lactic Acid Bacteria Cultures.
Fungal spoilage of dairy products is a major concern due to food waste and economical losses, some fungal metabolites may furthermore have adverse effects on human health. The use of lactic acid bacteria (LAB) is emerging as a potential clean label alternative to chemical preservatives. Here, our aim was to characterize the growth potential at three storage temperatures (5, 16, and 25°C) of a panel of molds (four Mucor and nine Penicillium strains) isolated from dairy products, then investigate the susceptibility of the molds toward 12 LAB cultures. Fungal cell growth and morphology in malt extract broth was monitored using oCelloScope at 25°C for 24 h. Mucor plumbeus 01180036 was the fastest growing and Penicillium roqueforti ISI4 (P. roqueforti ISI4) the slowest of the tested molds. On yogurt-agar plates, all molds grew at 5, 16, and 25°C in a temperature-dependent manner with Mucor strains growing faster than Penicillium strains regardless of temperature. The sensitivity toward 12 LAB cultures was tested using high-throughput overlay method and here all the molds except P. roqueforti ISI4 were strongly inhibited. The antifungal action of these LAB was confirmed when spotting mold spores on agar plates containing live cells of the LAB strains. However, if cells were removed from the fermentates, the inhibitory effects decreased markedly. The antifungal effects of volatiles tested in a plate-on-plate system without direct contact between mold and LAB culture media were modest. Some LAB binary combinations improved the antifungal activity against the growth of several molds beyond that of single cultures in yogurt serum. The role of competitive exclusion due to manganese depletion was examined as a possible antifungal mechanism for six Penicillium and two Mucor strains. It was shown that this mechanism was a major inhibition factor for the molds tested apart from the non-inhibited P. roqueforti ISI4 since addition of manganese with increasing concentrations of up to 0.1 mM resulted in partly or fully restored mold growth in yogurt. These findings help to understand the parameters influencing the mold spoilage of dairy products and the interactions between the contaminating strains, substrate, and bioprotective LAB cultures.
Shi C
，Knøchel S
《Frontiers in Microbiology》
Development of antifungal ingredients for dairy products: From in vitro screening to pilot scale application.
Biopreservation represents a complementary approach to traditional hurdle technologies for reducing microbial contaminants (pathogens and spoilers) in food. In the dairy industry that is concerned by fungal spoilage, biopreservation can also be an alternative to preservatives currently used (e.g. natamycin, potassium sorbate). The aim of this study was to develop antifungal fermentates derived from two dairy substrates using a sequential approach including an in vitro screening followed by an in situ validation. The in vitro screening of the antifungal activity of fermentates derivating from 430 lactic acid bacteria (LAB) (23 species), 70 propionibacteria (4 species) and 198 fungi (87 species) was performed against four major spoilage fungi (Penicillium commune, Mucor racemosus, Galactomyces geotrichum and Yarrowia lipolytica) using a cheese-mimicking model. The most active fermentates were obtained from Lactobacillus brevis, Lactobacillus buchneri, Lactobacillus casei/paracasei and Lactobacillus plantarum among the tested LAB, Propionibacterium jensenii among propionibacteria, and Mucor lanceolatus among the tested fungi. Then, for the 11 most active fermentates, culture conditions were optimized by varying incubation time and temperature in order to enhance their antifungal activity. Finally, the antifungal activity of 3 fermentates of interest obtained from Lactobacillus rhamnosus CIRM-BIA1952, P. jensenii CIRM-BIA1774 and M. lanceolatus UBOCC-A-109193 were evaluated in real dairy products (sour cream and semi-hard cheese) at a pilot-scale using challenge and durability tests. In parallel, the impact of these ingredients on organoleptic properties of the obtained products was also assessed. In semi-hard cheese, application of the selected fermentates on the cheese surface delayed the growth of spoilage molds for up to 21 days, without any effect on organoleptic properties, P. jensenii CIRM-BIA1774 fermentate being the most active. In sour cream, incorporation of the latter fermentate at 2 or 5% yielded a high antifungal activity but was detrimental to the product organoleptic properties. Determination of the concentration limit, compatible with product acceptability, showed that incorporation of this fermentate at 0.4% prevented growth of fungal contaminants in durability tests but had a more limited effect against M. racemosus and P. commune in challenge tests. To our knowledge, this is the first time that the workflow followed in this study, from in vitro screening using dairy matrix to scale-up in cheese and sour cream, is applied for production of natural ingredients relying on a large microbial diversity in terms of species and strains. This approach allowed obtaining several antifungal fermentates which are promising candidates for dairy products biopreservation.
Garnier L
，Mounier J
，Lê S
，Pawtowski A
，Pinon N
，Camier B
，Chatel M
，Garric G
，Thierry A
，Coton E
，Valence F
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Screening of Antifungal Lactic Acid Bacteria as Bioprotective Cultures in Yogurt and a Whey Beverage.
The demand for preservative-free food products is rising, and biopreservation is a potential alternative to replace or reduce the use of chemical preservatives. The objectives of this study were to assess the antifungal activity of lactic acid bacteria (LAB; n = 98) and the efficacy and applicability of the chosen bioprotective cultures against fungal spoilers in dairy products. First, 14 antifungal strains were preliminarily screened by in vitro tests against Pichia pastoris D3, Aspergillus niger D1, Geotrichum candidum N1, Kluyveromyces marxianus W1, and Penicillium chrysogenum B1 and validated by challenge tests in yogurt, indicating that the fungal-inhibiting activity of LAB was species specific and yogurt fermented with antifungal LAB cultures was more effective in extending shelf life. Second, the chosen 14 LAB strains were identified by the 16S rDNA sequence analysis and carbohydrate fermentation test. The results were as follows: nine strains were Lactobacillus plantarum, three were Lactobacillus paracasei, one was Enterococus faecium, and one was Lactobacillus rhamnosus. Among them, active L. plantarum N7 was the chosen and studied factor affecting antifungal activity by using the response surface methodology. Finally, in situ tests were conducted to validate the activity of L. plantarum N7 in actual dairy products (whey beverages). Physicochemical and microbial indices of whey beverages during storage indicated that antifungal L. plantarum N7 could slow yeast growth and be candidates of interest for industrial applications.
Xu R
，Sa R
，Jia J
，Li L
，Wang X
，Liu G
... -
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Evaluation of the efficacy of commercial protective cultures to inhibit mold and yeast in cottage cheese.
Biopreservation is defined as using microbes, their constituents, or both to control spoilage while satisfying consumer demand for clean-label products. The study objective was to investigate the efficacy of bacterial cultures in biopreserving cottage cheese against postprocessing fungal contamination. Cottage cheese curd and dressing were sourced from a manufacturer in New York State. Dressing was inoculated with 3 different commercial protective cultures-PC1 (mix of Lacticaseibacillus spp. and Lactiplantibacillus spp.), PC2 (Lacticaseibacillus rhamnosus), and PC3 (Lactic. rhamnosus)-following the manufacturer recommended dosage and then mixed with curd. A control with no protective culture was included. Nine species of yeast (Candida zeylanoides, Clavispora lusitaniae, Debaryomyces hansenii, Debaryomyces prosopidis, Kluyveromyces marxianus, Meyerozyma guilliermondii, Pichia fermentans, Rhodotorula mucilaginosa, and Torulaspora delbrueckii) and 11 species of mold (Aspergillus cibarius, Aureobasidium pullulans, Penicillium chrysogenum, Penicillium citrinum, Penicillium commune, Penicillium decumbens, Penicillium roqueforti, Mucor genevensis, Mucor racemosus, Phoma dimorpha, and Trichoderma amazonicum) were included in the study. Fungi strains were previously isolated from dairy processing environments and were inoculated onto the cheese surface at a rate of 20 cfu/g. Cheese was stored at 6 ± 2°C. Yeast levels were enumerated at 0, 7, 14, and 21 d postinoculation. Mold growth was visually observed on a weekly basis through d 42 of storage and imaged. Overall, the protective cultures were limited in their ability to delay the outgrowth in cottage cheese, with only 8 of the 20 fungal strains showing an effect of the cultures compared with the control. The protective cultures were not very effective against yeast, with only PC1 able to delay the outgrowth of 3 strains: D. hansenii, Tor. delbrueckii, and Mey. guilliermondii. The efficacy of these protective cultures against molds in cottage cheese was more promising, with all protective cultures showing the ability to delay spoilage of at least 1 mold strain. Both PC1 and PC2 were able to delay Pen. chrysogenum and Pho. dimorpha outgrowth, and PC1 also delayed Pen. commune, Pen. decumbens, and Pen. roqueforti to different extents compared with the controls. This study demonstrates that commercial lactic acid bacteria cultures vary in their performance to delay mold and yeast outgrowth, and thus each protective culture should be evaluated against the specific strains of fungi of concern within each specific dairy facility.
Makki GM
，Kozak SM
，Jencarelli KG
，Alcaine SD
... -
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ResearchGate
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