Freeze-dried Cultures Research Articles

Effects of adding autochthonous Lacticaseibacillus paracasei culture on the characteristics of Minas Frescal goat cheese and its survival under simulated gastrointestinal conditions

A freeze-dried lactic acid culture was prepared with an autochthonous strain of Lacticaseibacillus paracasei TRA061676 (Lp-TRA061676), cultivated in skimmed goat milk and its technological suitability in the production of Minas Frescal Goat Cheese (MFGC) was investigated. The viability of the freeze-dried culture was greater than 8.92 Log CFU g−1 during 360 days of storage at −22 °C. Lp-TRA061676 incorporated into MFGC showed remarkable survival, with populations of 9.65 Log CFU g−1 and a survival rate under in vitro gastrointestinal tract conditions of 96.90% for 28 days at 4 °C. Lp-TRA061676 did not modify the proximate composition of MFGC, but it affected the physicochemical characteristics and instrumental color, and improved the product's texture. These results demonstrate the suitability of Lp-TRA061676 for producing MFGC and the potential of this cheese to be used as an effective matrix to protect the strain from gastrointestinal conditions.

Efficiency of freeze- and spray-dried microbial preparation as active dried starter culture in kombucha fermentation.

Kombucha is a widely consumed fermented beverage produced by fermenting sweet tea with a symbiotic culture of bacteria and yeast (SCOBY). The dynamic nature of microbial communities in SCOBY may pose challenges to production scale-up due to unpredictable variations in microbial composition. Using identified starter strains is a novel strategy to control microorganism composition, thereby ensuring uniform fermentation quality across diverse batches. However, challenges persist in the cultivation and maintenance of these microbial strains. This study examined the potential of microencapsulated kombucha fermentation starter cultures, specifically Komagataeibacter saccharivorans, Levilactobacillus brevis and Saccharomyces cerevisiae, through spray-drying and freeze-drying. Maltodextrin and gum arabic-maltodextrin were employed as carrier agents. Our results revealed that both spray-dried and freeze-dried samples adhered to physicochemical criteria, with low moisture content (2.18-7.75%) and relatively high solubility (65.75-87.03%) which are appropriate for food application. Freeze-drying demonstrated greater effectiveness in preserving bacterial strain viability (88.30-90.21%) compared to spray drying (74.92-78.66%). Additionally, the freeze-dried starter strains demonstrated similar efficacy in facilitating kombucha fermentation, compared to the SCOBY group. The observations included pH reduction, acetic acid production, α-amylase inhibition and elevated total polyphenol and flavonoid content. Moreover, the biological activity, including antioxidant potential and in vitro tyrosinase inhibition activity, was enhanced in the same pattern. The freeze-dried strains exhibited consistent kombucha fermentation capabilities over a three-month preservation, regardless of storage temperature at 30 or 4 °C. These findings highlight the suitability of freeze-dried starter cultures for kombucha production, enable microbial composition control, mitigate contamination risks and ensure consistent product quality. © 2024 Society of Chemical Industry.

An effective strategy for improving the freeze-drying survival rate of Lactobacillus curvatus and its potential protective mechanism

Freeze-drying is a common method to prepare dried starter cultures with high stability and a long storage time. However, the bacterial cells suffer varying degrees of damage during the freeze-drying process. This study aimed to provide an effective strategy to improve the survival rate of Lactobacillus curvatus and to explore its potential protective mechanisms. The survival rates of bacterial cells in different cryoprotectants before and after freeze-drying were measured using the plate count method. The protective effects of composite cryoprotectants on bacterial cells were investigated from multiple perspectives, including key enzyme activities, the internal morphology of bacterial cells, and cell membrane integrity. Results showed that the freeze-drying survival rate of bacterial cells in the composite cryoprotectants consisting of trehalose, oleic acid and skimmed milk (85.38%) was significantly improved compared to that in PBS solution (2.04%). The addition of composite cryoprotectants could reduce damage to the cell wall and cell membrane, maintain the normal morphology of bacterial cells and cell membrane integrity, and improve key enzyme activities (lactate dehydrogenase, pyruvate kinase and ATPases) during the freeze-drying process. The viability of freeze-dried cultures was also monitored at different temperatures during storage, and the cell survival of freeze-dried cultures at −20 °C was the highest, reaching 61.13%. The findings obtained in this work offer a new method to improve the viability of Lactobacillus curvatus after freeze-drying, and may promote the wide use of freeze-dried cultures in the food fermentation industry.

Optimizing ruminant nutrition: Insights from a comprehensive analysis of silage composition and in vitro gas production dynamics using nonlinear models

Achieving sustainable livestock management necessitates optimizing animal production while minimizing environmental impact. To achieve this, feed efficiency must be enhanced, and nutrition blueprints must be understood. In ruminant nutrition, this is of paramount importance, as it exposes degradation kinetics and nutritional benchmarks, allowing feed management and formulations to be more ecologically balanced. Previous research efforts have focused on exploring the relationship between a restricted set of nutrient parameters and the in vitro gas production dynamics. In the current study, an extensive dataset derived from freeze-dried kefir culture treated white clover silage was used to examine intricate relationships between eight nonlinear models and diverse variables. This dataset contains in vitro gas production data along with nutritional composition, microbial populations, fermentation quality, digestibility, mineral concentration, and fatty acid profiles. Through rigorous application of mathematical models, the performance in capturing gas production dynamics was critically assessed. Among these, the Michaelis‒Menten (MM) and Mitscherlich (MIT) models fit the data well and offer superior predictions of gas production dynamics. Asymptotic gas volume was negatively correlated with crude protein content, emphasizing the influence of protein on gas production. Fiber composition plays a significant role in fermentation kinetics, as evidenced by significant correlations between degradation rate constant and crude protein concentrations. The degradation rate constant of insoluble fraction exhibited significant positive correlations with crude protein and neutral detergent fiber contents. Moreover, mineral content had significant effects on gas production dynamics. Zinc content showed a strong and significant positive correlation with the gas production rate coefficient, underscoring its crucial role in enhancing microbial activity. Conversely, calcium content displayed a significant but weak negative correlation with the final asymptotic gas volume, indicating its potential to modulate gas production. In summury, this study provides detailed insights into the intricate relationship between mathematical models and various variables in rumen fermentation. The MM and MIT models have proven to be robust tools, offering nuanced perspectives on gas production dynamics. These findings pave the path for improving sustainable ruminant nutritional practices and refining feed management strategies.

Impact of frequency and duration of freeze-dried inactivated tissue culture hepatitis A vaccine (Aimmugen®) vaccination on antibody titers; a japanese cross-sectional study

BackgroundThe effect of the timing of additional doses and the long-term persistence of lyophilized inactivated tissue culture hepatitis A (HA) vaccine (Aimmugen®) on antibodies is unknown. MethodsA single-center, cross-sectional, observational study was conducted in collaboration with the Japan Air Self-Defense Force, whose personnel were immunized with Aimmugen® when deployed to endemic areas. Patients who consented to this study after a medical examination with blood sampling between June 2022 and February 2023 were included; HA-IgG level in the residual serum was measured using the chemiluminescent immunoassay method. The exact vaccination history was investigated based on immunization records maintained by the Ministry of Defense, and a questionnaire was used to collect confounding factors. ResultsOf the 181 participants observed, 49 were in the unvaccinated group, and 132 were in the vaccinated group. Out of the vaccinated group, 6.8 % received either one or two doses, 40.9 % received three doses, and 52.3 % received more than four doses. IgG antibody titers (S/CO value) in each group (0, 1 or 2, 3, and over 4) increased in a frequency-dependent manner, with those vaccinated over four times showing significantly higher IgG antibody titers than all other groups (0.19 ± 0.10 vs 3.66 ± 3.00 vs 7.63 ± 3.57 vs 10.57 ± 1.86, respectively). When the number of months elapsed from the last vaccination to the date of blood collection in each group was plotted against IgG antibody titer, the slope of the regression line flattened out from a decreasing trend in the order 1 or 2, 3, over 4. ConclusionsThree doses of Aimmugen® are efficacious, but four or more doses induce more robust and sustained antibody production. Additionally, four or more doses may be effective when there is a need to ensure long-term immunity or risk of prolonged exposure.

An open-label, non-randomized study investigating the safety and efficacy of smallpox vaccine, LC16, as post-exposure prophylaxis for mpox

ABSTRACT Mpox is an acute exanthematous disease caused by the monkeypox virus. Since May 2022, it has spread as a community-acquired infection, mainly in Europe and the United States, and urgent measures to prevent this infection were also required in Japan. In this study, we investigated the post-exposure prophylaxis of mpox and safety after inoculating the smallpox vaccine. Participants in close contact with patients with mpox were inoculated with “Freeze-dried cell culture Smallpox Vaccine LC16,” within 14 days after close contact. Six cases were registered, and all the participants were inoculated. No mpox symptoms or related complications were observed in the participants for 21 days after the close contact. Adverse events due to inoculation, such as rash, fever, lymphadenopathy, and local reaction at the inoculation site (comprising erythema, swelling, induration, and pain) were observed in the participants; however, all inoculation-related events were non-severe and non-serious, and the participants recovered during the 28-day observation period. The findings of this study suggest that inoculation with LC16 is an effective post-exposure prophylaxis in individuals who had close contact with patients with mpox. Further large-scale studies are warranted to validate these findings.

Impact of media culture, freeze-drying and storage conditions on preservation of Lacticaseibacillus paracasei 90: Viability and metabolic potential as a secondary culture in semi-hard cheese

Freeze-drying is the main process used to preserve lactic bacterial cultures at an industrial level; however, their viability and metabolic activity can be affected not only by the process per se but also by storage conditions. This study investigated the effect of freeze-drying and long-term storage (14 months) on the viability and metabolic activity of Lacticaseibacillus paracasei 90 (L90) grown in three culture media. The freeze-drying process did not influence its culturability and viability, regardless of the culture media used. The storage of the freeze-dried cultures at room temperature had a negative impact on viability, culturability and its ability to grow and acidify milk. However, L90 was able to withstand a long period of storage at low temperatures (14 months, 4 °C) and keep the technological features desirable of a ripening culture. Overall, the results demonstrate the robustness of this strain as a secondary culture in the food industry.

The Enterococcus secretome inhibits the growth of Mycobacterium tuberculosis complex mycobacteria.

Enterococcus mundtii , a commensal intestinal bacterium, was demonstrated to inhibit the growth of some Mycobacterium tuberculosis complex (MTC) species that cause tuberculosis in humans and mammals. To further explore this preliminary observation, we cross-investigated five E. mundtii strains and seven MTC strains representative of four MTC species using a standardized quantitative agar well diffusion assay. All five E. mundtii strains, calibrated at 10 MacFarland, inhibited the growth of all M. tuberculosis strains with various susceptibility profiles, but no inhibition was observed with lower inoculums. Further, eight E. mundtii freeze-dried cell-free culture supernatants (CFCS) inhibited the growth of M. tuberculosis , Mycobacterium africanum, Mycobacterium bovis and Mycobacterium canettii, the most susceptible MTC species (inhibition diameter 25±1 mm), proportionally to CFCS protein concentrations. The data reported here indicate that the E. mundtii secretome inhibited growth of all MTC species of medical interest, which broadens previously reported data. In the gut, the E. mundtii secretome may modulate the expression of tuberculosis, exhibiting an anti-tuberculosis effect, with some protective roles in human and animal health.

Application of antifungal metabolites from Streptomyces philanthi RL-1-178 for maize grain coating formulations and their efficacy as biofungicide during storage

The major safety risk of maize grain is contamination with mycotoxins. In this study, a maize-coating formulation containing freeze-dried culture filtrate of Streptomyces philanthi RL-1-178 (DCF RL-1-178) was developed and evaluated to prevent the growth of mycotoxins during maize grain storage. In vitro studies using confrontation tests on PDA plates indicated that S. philanthi RL-1-178 inhibited the growth of Aspergillus parasiticus TISTR 3276 (89.0%) and A. flavus PSRDC-4 (95.0%). The maize grain coating formulations containing the DCF RL-1-178 (0, 5, 10, and 15% (v/v)) and the polymer polyvinylpyrrolidone (PVP-K90, 4.0% (w/v)) were tested for their efficacy in In vitro and during 5 months storage. In In vitro assay, maize coating formular containing the optimum concentration (15.0%, v/v) of the DCF RL-1-178 exhibited 54.80% and 54.17% inhibition on the growth of A. parasiticus TISTR 3276 and A. flavus PSRDC-4 respectively. The inhibition was also illustrated by the microstructures of interactions between the coated maize grains with or without the DCF RL-1-178 and the fungal pathogens observed under microscope and SEM. Incorporating the DCF RL-1-178 or fungicidal Metalaxyl® into the polymer PVP-K90 maize grains coating resulted in the complete inhibition of the production of aflatoxin B1 (analysed by HPLC) by the two aflatoxigenic pathogens after 5 months storage at room temperature. However, the shelf-life was shortened to only 3 months during storage at room temperature with 90% relative humidity. Overall, the application of the 10-15% DCF RL-1-178 into the maize grain coating formular provides a new alternative measure to control the mycotoxins during storage for at least 5 months. The In vitro cell cytotoxicity study showed that a concentration of 15% (v/v) or 1000μg/mL of the DCF RL-1-178 had a strong cytotoxic effect on Vero cells. These findings indicate that DCF RL-1-178 is a potential biofungicide for controlling mycotoxins contamination in maize seed storage for planting, but not maize grain storage for animal feed.

Long survival of Neisseria meningitidis in freeze-dried cultures maintained in potentially unsuitable conditions

Currently, diagnosis of Neisseria meningitidis in nasopharyngeal samples can be made by culture and nucleic acid amplification techniques.1 In Cuba, molecular diagnosis of meningococcal meningitis was introduced at the National Reference Laboratory for Neisseria (NRLN), in 2010, through a PCR that amplifies a fragment of the ctrA gene using the protocol described by Taha, 2000.2 This gene codes for a capsule protein that regulates the adhesion of N. meningitidis to the host, and 16 to 28% of meningococci isolates, especially in the nasopharynx, lack this gene.2 In contrast, the sodC gene, related to the production of superoxide-dismutase of this organism, is less sensitive to antigenic variation, hence its importance for molecular diagnosis in patients and asymptomatic carriers.3 Information on the meningococcal carriage is essential for public health policy,4. Still, the high number of invasive meningococci disease (IMD) affecting Cuba during the 1980s and the absence of molecular tools prevented its accurate microbiological diagnosis in carriers.

A Comparative Study of Skimmed Milk and Cassava Flour on the Viability of Freeze-Dried Lactic Acid Bacteria as Starter Cultures for Yogurt Fermentation.

The purpose of this study was to evaluate the survival rates and fermentation performance of three freeze-dried lactic acid bacterial cultures previously isolated from Ghanaian traditional fermented milk. LAB cultures, i.e., Lactobacillus delbrueckii, Lactococcus lactis and Leuconostoc mesenteroides, were frozen in the chamber of a Telstar (Lyoquest) laboratory freeze dryer for 10 h at -55 °C (as single and combined cultures) using skimmed milk and cassava flour as cryoprotectants held in plastic or glass cryovials. For viability during storage, freeze-dried LAB cultures were stored in a refrigerator (4 °C) and at room temperature (25 °C) for 4 weeks. The survival of freeze-dried cultures was determined by growth kinetics at 600 nm (OD600). The performance of freeze-dried LAB cultures after 4 weeks of storage was determined by their growth, acidification of milk during yogurt fermentation and consumer sensory evaluation of fermented milk using a nine-point hedonic scale. The survival rates for LAB ranged between 60.11% and 95.4% following freeze-drying. For single cultures, the highest survival was recorded for Lactobacillus delbrueckii (L12), whereas for combined cultures, the highest survival was observed for Lactococcus lactis (L3) combined with Lactobacillus delbrueckii (L12). The consumer acceptability results showed that yogurts produced from a combined starter culture of Lactococcus lactis and Lactobacillus delbrueckii or from a single culture of Lactococcus lactis were the most preferred products with Lactococcus lactis and Lactobacillus delbrueckii possessing high survival rates and high consumer acceptability in yogurt production. These findings are crucial and can be adopted for large-scale production and commercialization of yogurt.

Use of commercial protective cultures in portioned sheep milk cheeses to control Listeria monocytogenes.

The main objective of this study was to innovate soft and semi-cooked sheep milk cheese production processes with the use of a commercial protective culture able to control Listeria monocytogenes growth. A freeze-dried commercial culture of Lactobacillus plantarum was tested in DS cheese and PS cheese, two types of pasteurized sheep milk, raw-paste cheeses aged for no less than 20 and 30 days respectively. In the first step, in vitro tests were conducted to identify the most suitable matrix for the growth of L. plantarum in order to create a subculture that could be used at industrial cheese-making plants. During the second phase of the study, L. plantarum culture was introduced in the manufacturing process of the cheeses in a production plant. Finally, a challenge test was conducted on portioned DS and PS cheeses in order to evaluate the activity of the protective culture against L. monocytogenes: the cheeses were portioned, experimentally contaminated with L. monocytogenes strains, vacuum packed and stored at +4°C (correct storage conditions) and at +10°C (thermal abuse). Cheeses were analysed at the end of the shelf-life to evaluate the presence and growth of L. monocytogenes, to enumerate lactic acid bacteria and determine chemicalphysical features. The results confirmed that protective cultures are a useful technological innovation to control L. monocytogenes growth during cheese storage without altering composition, microflora and chemical- physical characteristics of the product. However, the use of protective cultures should be applied as an integration of risk control measures and not as a substitute for preventive actions.

Novel Functional Grape Juices Fortified with Free or Immobilized Lacticaseibacillus rhamnosus OLXAL-1.

During the last decade, a rising interest in novel functional products containing probiotic microorganisms has been witnessed. As food processing and storage usually lead to a reduction of cell viability, freeze-dried cultures and immobilization are usually recommended in order to maintain adequate loads and deliver health benefits. In this study, freeze-dried (free and immobilized on apple pieces) Lacticaseibacillus rhamnosus OLXAL-1 cells were used to fortify grape juice. Juice storage at ambient temperature resulted in significantly higher (>7 log cfu/g) levels of immobilized L. rhamnosus cells compared to free cells after 4 days. On the other hand, refrigerated storage resulted in cell loads > 7 log cfu/g for both free and immobilized cells for up to 10 days, achieving populations > 109 cfu per share, with no spoilage noticed. The possible resistance of the novel fortified juice products to microbial spoilage (after deliberate spiking with Saccharomyces cerevisiae or Aspergillus niger) was also investigated. Significant growth limitation of both food-spoilage microorganisms was observed (both at 20 and 4 °C) when immobilized cells were contained compared to the unfortified juice. Keynote volatile compounds derived from the juice and the immobilization carrier were detected in all products by HS-SPME GC/MS analysis. PCA revealed that both the nature of the freeze-dried cells (free or immobilized), as well as storage temperature affected significantly the content of minor volatiles detected and resulted in significant differences in the total volatile concentration. Juices with freeze-dried immobilized cells were distinguished by the tasters and perceived as highly novel. Notably, all fortified juice products were accepted during the preliminary sensory evaluation.

Developing Stable Freeze-Dried Functional Ingredients Containing Wild-Type Presumptive Probiotic Strains for Food Systems

Designing stable dried functional food ingredients and foods containing live probiotic cultures maintaining high viable cell loads at the time of consumption is a challenge for the industry. The aim of the present study was the development of stable freeze-dried functional food ingredients with enhanced shelf-life during long storage. Zea flakes, pistachios, and raisins were used as immobilization supports for the wild-type presumptive probiotic strains Pediococcus acidilactici SK and Lactiplantibacillus plantarum F4, while L. plantarum B282 was used as a reference strain. Cell survival was monitored during storage at room and refrigerated temperatures for up to 6 months. Levels of freeze-dried cultures were maintained up to 7.2 logcfu/g after 6 months storage at room temperature and up to 8.5 logcfu/g at refrigerator temperature, in contrast to free cell levels that ranged <7 logcfu/mL, suggesting the positive effects of immobilization and freeze-drying on cell viability. Of note, levels of freeze-dried immobilized P. acidilactici SK cells on zea flakes and pistachios remained stable after 6 months of storage at 4 °C, ranging 8.1–8.5 logcfu/g (survival rates 98.2 and 99.7%, respectively). The technology developed presents important advantages for the maintenance of cell viability during storage, assuring stability of ready-to-use functional food ingredients that could be directly incorporated in food systems.

Quality assessment of buffalo milk Chakka prepared from different starter culture

Chakka is a soured dairy product that is producedby removing the whey from fermented milk (dahi, known as In-dian yogurt). The starter culture plays a significant role in deter-mining the overall elemental quality of fermented dairy products. The present study was carried to find out the suitability of three different cultures for development of chakka viz. T1 (1.5% Mixed microbial culture), T2 (1.5% bacterial culture, STI-13 freeze-dried Lactic culture), and T3 (1.5% yeast cultures). Among the different culture various quality characteristics, physico-chemical param-eters and sensory characteristics were assessed for their ability to form chakka. Curd prepared from the Lactobacillus culture(T2) was found most suitable for preparation of chakka as T2delivered highest yield of curd mass, protein, least pH value and moisture content compared to microbial culture and yeast culture. Sensory attributes of chakka also varied significantly (P0.05) among cultures. All sensory attributes rated higher for T2(Lactobacillus cultures). Therefore, result of the study concluded that T2 (1.5% Lactobacillus) culture was most appropriate for formulation of chakka.

Antifungal and antiaflatoxigenic mechanism activity of freeze-dried culture filtrate ofStreptomyces philanthiRL-1-178 on the two aflatoxigenic fungi and identification of its active component

The study reports the antifungal and antiaflatoxigenic mechanism activity of freeze-dried culture filtrate of Streptomyces philanthi RL-1-178 (DCF RL-1-178) against two aflatoxigenic strains (Aspergillus parasiticus and A. flavus) and identification of its active component. Significant inhibition in ergosterol biosynthesis by the DCF RL-1-178 appeared on the plasma membrane. Moreover, the DCF RL-1-178 showed dose-dependent inhibition of methylglyoxal (MG) (an aflatoxin inducer) biosynthesis and exhibited a novel antiaflatoxigenic action mechanism. Significant impairments in enzymatic [superoxide dismutase (SOD) and catalase (CAT)] and nonenzymatic [oxidized and reduced glutathione (GSH) and ratio of oxidized and reduced glutathione (GSSG)] anti-oxidative defense molecules were observed in the two aflatoxigenic cells. The active component of the DCF RL-1-178 was identified as natamycin. The natamycin exhibited against A. parasiticus and A. flavus with the minimum inhibitory concentration (MIC) values of 0.5 and 1.0 µg ml-1, respectively, while the minimum fungicidal concentration values were the same (4.0 µg ml-1). The DCF RL-1-178 containing natamycin exhibited the following effects: (1) inhibition of cellular ergosterol biosynthesis on plasma membrane, (2) reduction in MG (aflatoxin inducer) confirmed novel antiaflatoxigenic mechanism of action, and (3) caused remarkable debasement in antioxidant defense enzymes (SOD and CAT) and nonenzymatic defense molecules (GSH and GSSG) revealing biochemical mechanism of action.

Enhanced virulence of Beauveria bassiana against Thrips tabaci by the addition of bacterial metabolites derived from Xenorhabdus hominickii

• Immunosuppressive agents are supposed to enhance the virulence of microbial pathogens against insect pests. • The culture broth of Xenorhabdus hominickii inhibited immune responses of Thrips tabaci . • The addition of the bacterial broth to Beauveria bassiana enhanced the fungal insecticidal activity. Xenorhabdus and Photorhabdus are two bacterial genera specifically symbiotic to Steinernema and Heterorhabditis , which are the entomopathogenic nematode genera, respectively. These bacteria are well known to produce potent secondary metabolites suppressing insect immune responses. This study aimed to develop a potent microbial insecticide against the onion thrips, Thrips tabaci , using the bacterial metabolites. Among the chemical insecticides that have been used to control the thrips, spinosad was highly effective against both larvae and adults of T. tabaci . Three different entomopathogenic fungi were also effective to kill the thrips. However, the fungal virulence was much less than the control efficacy of the chemical insecticide, spinosad. To enhance the fungal virulence of Beauveria bassiana (Bb), the bacterial culture broth of Xenorhabdus / Photorhabdus was added to suppress the thrips immune defense. Among six different bacterial species, X. hominickii (Xh) produced highly potent metabolites to enhance the fungal virulence. Indeed, four different bacterial metabolites (GameXPeptide, benzylideneacetone, oxindole, and 3-ethoxy-4-methoxyphenol) of the bacteria suppressed the gene expressions of an antimicrobial peptide, lysozyme , which was highly inducible to the fungal infection. To optimize the mixture ratio of fungal and bacterial pathogens, the fungal conidia and bacterial culture broth were freeze-dried and mixed in different ratios. Laboratory and field assays showed that a mixture spray of freeze-dried Xh culture broth (3 g) and Bb conidia (1.17 × 10 9 conidia) in a liter was effective to control T. tabaci infesting welsh onion.

Efficacy of the antifungal metabolites of Streptomyces philanthi RL-1-178 on aflatoxin degradation with its application to prevent aflatoxigenic fungi in stored maize grains and identification of the bioactive compound.

Aflatoxin B1 is a potent carcinogen produced by Aspergillus flavus (A. flavus) and Aspergillus. parasiticus (A. parasiticus), mainly during grain storage. The efficacy of the freeze-dried culture filtrate of Streptomyces philanthi (S. philanthi) strain RL-1-178 (DCF) on degradation of aflatoxin B1 (AFB1) were evaluated and its bioactive compounds were identified. The DCF at a concentration of 9.0% (w/v) completely inhibited growth and AFB1 production of A. parasiticus TISTR 3276 and A. flavus PSRDC-4 after 7days tested in yeast-extract sucrose (YES) medium and on stored maize grains after 28 and 14days incubation, respectively. This indicated the more tolerance of A. parasiticus over A. flavus. The DCF and bacterial cells of S. philanthi were capable to degrade AFB1 by 85.0% and 100% for 72h and 8days, respectively. This confirmed the higher efficacy of the DCF over the cells. After separation of the DCF on thin-layer chromatography (TLC) plate by bioautography bioassay, each active band was identified by liquid chromatography-quadrupole time of flight mass spectrometer (LC-Q-TOF MS/MS). The results revealed two compounds which were identified as azithromycin and an unknown based on mass ions of both ESI+ and ESI- modes. The antifungal metabolites in the culture filtrate of S. philanthi were proved to degrade aflatoxin B1. It could be concluded that the DCF may be applied to prevent the growth of the two aflatoxin-producing fungi as well as the occurrence of aflatoxin in the stored maize grains.

Probiotic Property and Anti-Obesity Effect of Lactiplantibacillus plantarum KC3.

Lactic acid bacteria are representative probiotics that have beneficial effects on humans. Nineteen strains among the 167 single strains from kimchi was selected and their physiological features were investigated. The selection of a strain was based on strong enzyme (lipase, α-amylase, and α-glucosidase) inhibitory activities and anti-obesity effects in the adipocytes. For the final selection, the strain Lactiplantibacillus plantarum KC3 was tested for its potential as a starter. To assess its functionality, a freeze-dried culture of L. plantarum KC3 was administered to a diet-induced obese mouse model receiving a high-fat diet. The animal group administered with L. plantarum KC3 showed significant body weight loss during the 12-week feeding period compared to the high-fat control group. This study investigated the physiological characteristics of selected strain and evaluated its potential as an anti-obesity probiotic in mice.

Improvement of Approaches to the Verification of the Vaccine Strain <i>Francisella tularensis</i> 15 NIIEG during Long-Term Storage

The aim of the study was to improve the methods for verifying the vaccine strain Francisella tularensis 15 NIIEG during long-term storage under current conditions.Materials and methods. The paper summarizes the results of studying the phenotypic and genetic properties of lyophilized cultures of the vaccine strain F. tularensis 15 NIIEG (1953, 1966, 1969, 1987, 1990, 2003, 2012 and 2013) stored at SCEMAP for a period of one to 60 years.Results and discussion. Previous studies have revealed that freeze-dried cultures of F. tularensis 15 NIIEG generally had the characteristics of the vaccine strain, with the exception of deviations from the regulatory requirements for residual virulence and specific safety. The stability of preservation of deletions in the pilA and pilE genes (the region of differentiation RD19) and the genes encoding lpp lipoprotein (RD18) in the vaccine strain, which was stored for various periods of time in a lyophilized state, has been confirmed. The vaccine-strain-specific mutation C178404T (by the genome of F. tularensis LVS strain, GenBank NCBI no. CP009694) has been identified, and an approach to determine it has been developed. The data obtained are promising as regards using the above deletions in the RD18/RD19 regions in combination with the C178404T mutation to assess the authenticity of the vaccine strain using molecular genetic methods. Thus, the conducted retrospective analysis of the data on the cultures of tularemia microbe vaccine strain from the 1940s to 2013 and the gathered experimental data, made it possible to supplement the uniform requirements for the manufacture, study, maintenance, storage and movement of F. tularensis 15 NIIEG vaccine strain with new evidence. Based on the results obtained, the authors have drawn a draft methodological recommendations of the federal level “Vaccinal strain Francisella tularensis 15 NIIEG: order of handling”.