Phenotypic and genotypic characterization of antimicrobial resistances reveals the effect of the production chain in reducing resistant lactic acid bacteria in an artisanal raw ewe milk PDO cheese.

Antimicrobial resistance (AMR) is a significant public health threat, with the food production chain, and, specifically, fermented products, as a potential vehicle for dissemination. However, information about dairy products, especially raw ewe milk cheeses, is limited. The present study analysed, for the first time, the occurrence of AMRs related to lactic acid bacteria (LAB) along a raw ewe milk cheese production chain for the most common antimicrobial agents used on farms (dihydrostreptomycin, benzylpenicillin, amoxicillin and polymyxin B). More than 200 LAB isolates were obtained and identified by Sanger sequencing (V1-V3 16S rRNA regions); these isolates included 8 LAB genera and 21 species. Significant differences in LAB composition were observed throughout the production chain (P ≤ 0.001), with Enterococcus (e.g., E. hirae and E. faecalis) and Bacillus (e.g., B. thuringiensis and B. cereus) predominating in ovine faeces and raw ewe milk, respectively, along with Lactococcus (L. lactis) in whey and fresh cheeses, while Lactobacillus and Lacticaseibacillus species (e.g., Lactobacillus sp. and L. paracasei) prevailed in ripened cheeses. Phenotypically, by broth microdilution, Lactococcus, Enterococcus and Bacillus species presented the greatest resistance rates (on average, 78.2 %, 56.8 % and 53.4 %, respectively), specifically against polymyxin B, and were more susceptible to dihydrostreptomycin. Conversely, Lacticaseibacillus and Lactobacillus were more susceptible to all antimicrobials tested (31.4 % and 39.1 %, respectively). Thus, resistance patterns and multidrug resistance were reduced along the production chain (P ≤ 0.05). Genotypically, through HT-qPCR, 31 antimicrobial resistance genes (ARGs) and 6 mobile genetic elements (MGEs) were detected, predominating Str, StrB and aadA-01, related to aminoglycoside resistance, and the transposons tnpA-02 and tnpA-01. In general, a significant reduction in ARGs and MGEs abundances was also observed throughout the production chain (P ≤ 0.001). The current findings indicate that LAB dynamics throughout the raw ewe milk cheese production chain facilitated a reduction in AMRs, which has not been reported to date.

Novel insight into the formation and improvement mechanism of physical property in fermented tilapia sausage by cooperative fermentation of newly isolated lactic acid bacteria based on microbial contribution.

Single starter can hardly elevate the gel property of fermented freshwater fish sausage. In this work, in order to improve the physical properties of tilapia sausage, two newly isolated strains of lactic acid bacteria (LAB), Latilactobacillus sakei and Pediococcus acidilactici were used for cooperative fermentation of tilapia sausage, followed by the revelation of their formation mechanisms during cooperative fermentation and their improvement mechanisms after comparison with natural fermentation. Both strains, especially L. sakei possessed good growth, acidification ability, and salt tolerance. The gel strength, hardness, springiness, chewiness, whiteness, acidification, and total plate count significantly elevated during cooperative fermentation with starters. Pediococcus, Acinetobacter, and Macrococcus were abundant before fermentation, while Latilactobacillus quickly occupied the dominant position after fermentation for 18-45 h with the relative abundance over 51.5 %. The influence of each genus on the physical properties was calculated through the time-dimension and group-dimension correlation networks. The results suggested that the increase of Latilactobacillus due to the good growth and metabolism of L. sakei contributed the most to the formation and improvement of gel strength, texture properties, color, acidification, and food safety of tilapia sausage after cooperative fermentation. This study provides a novel analysis method to quantitatively evaluate the microbial contribution on the changes of various properties. The cooperative fermentation of LAB can be used for tilapia sausage fermentation to improve its physical properties.

Study protocol: The role of milk matrix lipids in programming the immunoreactivity of proteins derived from lactic acid bacteria.

Food allergy is widely recognized as a significant health issue, having escalated into a global epidemic, subsequently giving rise to the development of numerous additional complications. Currently, the sole efficient method to curb the progression of allergy is through the implementation of an elimination diet. The increasing number of newly identified allergens makes it harder to completely remove or avoid them effectively. The immunoreactivity of proteins of bacterial origin remains an unexplored topic. Despite the substantial consumption of microbial proteins in our diets, the immunologic mechanisms they might induce require thorough validation. This stands as the primary objective of this study. The primary objective of this study was to evaluate the effects of bacterial proteins on the intestinal barrier and immune system parameters during hypersensitivity induction in both developing and mature organisms. The secondary objective was to evaluate the role of lipids in the immunoreactivity programming of these bacterial proteins. Notably, in this complex, comprehensively designed in vitro, in vivo, and ex vivo trial, the immunoreactivity of various bacterial proteins will be examined. In summary, the proposed study intends to address the knowledge gaps regarding the effects of Lactobacillus microbial proteins on inflammation, apoptosis, autophagy, and intestinal barrier integrity in a single study.

The predominant lactic acid bacteria and yeasts involved in the spontaneous fermentation of millet during the production of the traditional porridge Hausa koko in Ghana.

Spontaneous fermentation of cereals like millet involves a diverse population of microbes from various sources, including raw materials, processing equipment, fermenting receptacles, and the environment. Here, we present data on the predominant microbial species and their succession at each stage of the Hausa koko production process from five regions of Ghana. The isolates were enumerated using selective media, purified, and phenotypically characterised. The LAB isolates were further characterised by 16S rRNA Sanger sequencing, typed using (GTG)5 repetitive-PCR, and whole genome sequencing, while 28S rRNA Sanger sequencing was performed for yeast identification. The pH of the millet grains ranged from mean values of 6.02-6.53 to 3.51-3.99 in the final product, depending on the processors. The mean LAB and yeast counts increased during fermentation then fell to final counts of log 2.77-3.95 CFU/g for LAB and log 2.10-2.98 CFU/g for yeast in Hausa koko samples. At the various processing stages, the counts of LAB and yeast revealed significant variations (p < 0.0001). The species of LAB identified in this study were Limosilactobacillus pontis, Pediococcus acidilactici, Limosilactobacillus fermentum, Limosilactobacillus reuteri, Pediococcus pentosaceus, Lacticaseibacillus paracasei, Lactiplantibacillus plantarum, Schleiferilactobacillus harbinensis, and Weissella confusa. The yeasts were Saccharomyces cf. cerevisiae/paradoxus, Saccharomyces cerevisiae, Pichia kudriavzevii, Clavispora lusitaniae and Candida tropicalis. The identification and sequencing of these novel isolates and how they change during the fermentation process will pave the way for future controlled fermentation, safer starter cultures, and identifying optimal stages for starter culture addition or nutritional interventions. These LAB and yeast species are linked to many indigenous African fermented foods, potentially acting as probiotics in some cases. This result serves as the basis for further studies into the technological and probiotic potential of these Hausa koko microorganisms.

Antifungal mechanisms of binary combinations of volatile organic compounds produced by lactic acid bacteria strains against Aspergillusflavus.

Aspergillus flavus（A. flavus）, a common humic fungus known for its ability to infect agricultural products, served as the subject of investigation in this study. The primary objective was to assess the antifungal efficacy and underlying mechanisms of binary combinations of five volatile organic compounds (VOCs) produced by lactic acid bacteria, specifically in their inhibition of A. flavus. This assessment was conducted through a comprehensive analysis, involving biochemical characterization and transcriptomic scrutiny. The results showed that VOCs induce notable morphological abnormalities in A. flavus conidia and hyphae. Furthermore, they disrupt the integrity of the fungal cell membrane and cell wall, resulting in the leakage of intracellular contents and an increase in extracellular electrical conductivity. In terms of cellular components, VOC exposure led to an elevation in malondialdehyde content while concurrently inhibiting the levels of total lipids, ergosterol, soluble proteins, and reducing sugars. Additionally, the impact of VOCs on A. flavus energy metabolism was evident, with significant inhibition observed in the activities of key enzymes, such as Na+/K+-ATPase, malate dehydrogenase, succinate dehydrogenase, and chitinase. And they were able to inhibit aflatoxin B1 synthesis. The transcriptomic analysis offered further insights, highlighting that differentially expressed genes (DEGs) were predominantly associated with membrane functionality and enriched in pathways about carbohydrate and amino acid metabolism. Notably, DEGs linked to cellular components and energy-related mechanisms exhibited down-regulation, thereby corroborating the findings from the biochemical analyses. In summary, these results elucidate the principal antifungal mechanisms of VOCs, which encompass the disruption of cell membrane integrity and interference with carbohydrate and amino acid metabolism in A. flavus.

Lactic acid fermented microalgae and cyanobacteria as a new source of lipid reducing compounds: assessment through zebrafish Nile red fat metabolism assay and untargeted metabolomics.

Obesity is one of the most important threats to human health. Besides existing pharmacological or clinical interventions, novel effective and largely available solutions are still necessary. Among diverse natural resources, microalgae are well known for their complexity in the production of novel secondary metabolites. At the same time, lactic acid bacteria (LAB) are known for their capacity to metabolize, through fermentation, different matrices, and consequently to modify or produce new compounds with potential bioactivity. This work aimed to study the production of fermented microalgae and cyanobacteria, and to analyse their extracts in the zebrafish Nile red fat metabolism assay. Three microalgal species (Chlorella vulgaris, Chlorococcum sp. and Arthrospira platensis) were fermented with seven strains of LAB from 4 species (Lacticaseibacillus rhamnosus, Lacticaseibacillus casei, Lactobacillus delbrueckii bulgaricus and Lacticaseibacillus paracasei), derived from the UPCCO - University of Parma Culture Collection, Parma, Italy). All the selected strains were able to ferment the selected species of microalgae, and the most suitable substrate for LAB growth was Arthrospira platensis. Extracts from fermented Chlorella vulgaris and Chlorococcum sp. reduced significantly the neutral lipid reservoirs, which was not observed without fermentations. The strongest lipid reducing effect was obtained with Arthrospira platensis fermented with Lactobacillus delbrueckii bulgaricus 1932. Untargeted metabolomics identified some compound families, which could be related to the observed bioactivity, namely fatty acids, fatty amides, triterpene saponins, chlorophyll derivatives and purine nucleotides. This work opens up the possibility of developing novel functional foods or food supplements based on microalgae, since lactic acid fermentation enhanced the production of bioactive compounds with lipid reducing activities.

Identification of microorganisms in wet coffee fermentation Coffea arabica Var Catimor and Castillo in Jardín, Antioquia-Colombia, using culture-dependent methods / Identificación de microorganismos en fermentación húmeda de café Coffea arabica Var Catimor y Castillo en Jardín, Antioquia-Colombia, usando métodos cultivo-dependientes

Background: Mild Colombian coffees are recognized worldwide for their high-quality coffee cup. However, there have been some failures in post-harvest practices, such as coffee grain fermentation. These failures could occasionally lead to defects and inconsistencies in quality products and economic losses for coffee farmers. In Colombia, one of the fermentation methods most used by coffee growers is wet fermentation, conducted by submerging the de-pulped coffee beans for enough time in water tanks to remove the mucilage. Objectives: We evaluated the effect of the water (g)/de-pulped coffee (g) ratio (I: 0/25, II: 10/25, III: 20/25) and final fermentation time (24, 48, and 72 hours) on the total number of microbial groups. We also identified microorganisms of interest as starter cultures. Methods: We used a completely randomized experimental design with two factors; the effect of the water (g)/de-pulped coffee (g) ratio (I: 0/25, II: 10/25, III: 20/25) and final fermentation time (24, 48, and 72 hours), for 9 treatments with two replicates. During the coffee fermentation (1,950 g), the pH and °Brix were monitored. Total counts of different microbial groups (mesophiles, coliforms, lactic-acid bacteria, acetic-acid bacteria, and yeasts) were performed. Various isolates of microorganisms of interest as starter cultures (lactic-acid bacteria and yeasts) were identified using molecular sequencing techniques. Results: 21 lactic-acid bacteria (LAB) isolates and 22 yeasts were obtained from the different mini-batch fermentation systems. The most abundant lactic-acid bacteria species found were Lactiplantibacillus plantarum (46%) and Levilactobacillus brevis (31%). Pichia kluivery (39%) and Torulaspora delbrueckii (22%) were the most abundant yeast species. Conclusion The studied factors did not have effect over the microorganism's development. The identified bacterial and yeasts species have potential as starter cultures for better-quality coffees and in fermentation-related applications.

Antecedentes: Los cafés suaves lavados colombianos son reconocidos a nivel mundial por su buena puntuación sensorial; sin embargo, se han detectado fallas en las prácticas de postcosecha, como lo es la fermentación de los granos de café. Dichas fallas pueden causar defectos y carecer de consistencia en la calidad del producto, ocasionando pérdidas económicas para los caficultores. En Colombia, uno de los métodos más usados por los caficultores es la fermentación húmeda, la cual consiste en sumergir los granos de café despulpado en tanques con agua por un período de tiempo que permita la remoción del mucílago. Objetivos: La presente investigación evaluó la incidencia que tienen la proporción agua/granos despulpados de café (I: 0/25, II: 10/25, III: 20/25) y el tiempo final de fermentación (24, 48 y 72 horas) en el recuento final de grupos microbianos. Por otra parte, se identificaron taxonómicamente microorganismos de interés para su uso como cultivos iniciadores. Métodos: Mini-lotes consistieron en café despulpado (1950 g) puesto en recipientes de plástico abiertos y sumergidos en agua. Se aplicó un diseño experimental completamente aleatorizado de dos factores (proporción agua/ granos de café despulpado y tiempo) a tres niveles, para un total de nueve tratamientos con dos replicas. Durante las fermentaciones de café (1,950 g), el pH y los grados ºBrix, fueron monitoreados. Se realizaron los recuentos totales de los diferentes grupos microbianos: mesófilos, coliformes, bacterias ácido-lácticas, bacterias ácido-acéticas y levaduras. Se identificaron molecularmente diferentes aislados con potencial para ser usados como cultivos iniciadores (bacterias ácido-lácticas y levaduras). Resultados: Los resultados obtenidos mostraron que no hubo diferencia estádisticamente significativa entre los tratamientos aplicados y el recuento final de microorganismos. Un total de 21 aislados de bacterias ácido-lácticas (BAL) y 22 levaduras lograron obtenerse a partir de los diferentes sistemas de fermentación en mini-lote. Las especies de bacterias ácido-lácticas con mayor porcentaje acorde a su identificación taxonómica, corresponden a Lactiplantibacillus plantarum (46%), Levilactobacillus brevis (31%). Las especies de levaduras con mayor porcentaje acorde a su identificación taxonómica corresponden a Pichia kluivery (39%) y Torulaspora delbrueckii (22%). Conclusión Los factores estudiados no afectaron el crecimiento de ninguno de los grupos microbianos presentes en la fermentacion del café. Las especies de microorganismos identificados tienen potencial para se usados como cultivos starter o en aplicaciones dentro de las ciencias de fermentación.

New Genetic Determinants for qPCR Identification and the Enumeration of Selected Lactic Acid Bacteria in Raw-Milk Cheese.

Lactic acid bacteria (LAB) play an important role in the ripening of cheeses and contribute to the development of the desired profile of aroma and flavor compounds. Therefore, it is very important to monitor the dynamics of bacterial proliferation in order to obtain an accurate and reliable number of their cells at each stage of cheese ripening. This work aimed to identify and conduct a quantitative assessment of the selected species of autochthonous lactic acid bacteria from raw cow's milk cheese by the development of primers and probe pairs based on the uniqueness of the genetic determinants with which the target microorganisms can be identified. For that purpose, we applied real-time quantitative PCR (qPCR) protocols to quantify Lactobacillus delbrueckii subsp. bulgaricus, Streptococcus thermophilus, and Lactococcus lactis subsp. cremoris cells in cheese directly after production and over three-month and six-month ripening periods. While L. lactis subsp. cremoris shows good acidification ability and the ability to produce antimicrobial compounds, L. delbrueckii subsp. bulgaricus has good proteolytic ability and produces exo-polysaccharides, and S. thermophilus takes part in the formation of the diacetyl flavor compound by metabolizing citrate to develop aroma, they all play an important role in the cheese ripening. The proposed qPCR protocols are very sensitive and reliable methods for a precise enumeration of L. delbrueckii subsp. bulgaricus, S. thermophilus, and L. lactis subsp. cremoris in cheese samples.

The oral cavity and intestinal microbiome in children with functional constipation.

Constipation is a widespread problem in paediatric practice, affecting almost 30% of children. One of the key causal factors of constipation may be disturbances in the homeostasis of the gastrointestinal microbiome. The aim of the study was to determine whether the oral and fecal microbiomes differ between children with and without constipation. A total of 91 children over three years of age were included in the study. Of these, 57 were qualified to a group with constipation, and 34 to a group without. The saliva and stool microbiomes were evaluated using 16S rRNA gene amplicon sequencing. Functional constipation was associated with characteristic bacterial taxa in the fecal microbiota. Statistically significant differences were found at the family level: Burkholderiaceae (q = 0.047), Christensenellaceae (q = 0.047), Chlostridiaceae (q = 0.047) were significantly less abundant in the constipation group, while the Tannerellaceae (q = 0.007) were more abundant. At the genus level, the significant differences were observed for rare genera, including Christensenellaceae r-7 (q = 2.88 × 10-2), Fusicatenibacter (q = 2.88 × 10-2), Parabacteroides (q = 1.63 × 10-2), Romboutsia (q = 3.19 × 10-2) and Subdoligranulum (q = 1.17 × 10-2). All of them were less abundant in children with constipation. With the exception of significant taxonomic changes affecting only feces, no differences were found in the alpha and beta diversity of feces and saliva. Children with functional constipation demonstrated significant differences in the abundance of specific bacteria in the stool microbiome compared to healthy children. It is possible that the rare genera identified in our study which were less abundant in the constipated patients (Christensellaceae r-7, Fusicatenibacter, Parabacteroides, Romboutsia and Subdoligranulum) may play a role in protection against constipation. No significant differences were observed between the two groups with regard to the saliva microbiome.

Legume protein fermented by lactic acid bacteria: Specific enzymatic hydrolysis, protein composition, structure, and functional properties.

In recent years, with increasing emphasis on healthy, green, and sustainable consumption concepts, plant-based foods have gained popularity among consumers. As widely sourced plant-based raw materials, legume proteins are considered sustainable and renewable alternatives to animal proteins. However, legume proteins have limited functional properties, which hinder their application in food products. LAB fermentation is a relatively natural processing method that is safer than chemical/physical modification methods and can enrich the functional properties of legume proteins through biodegradation and modification. Therefore, changes in legume protein composition, structure, and functional properties and their related mechanisms during LAB fermentation are described. In addition, the specific enzymatic hydrolysis mechanisms of different LAB proteolytic systems on legume proteins are also focused in this review. The unique proteolytic systems of different LAB induce specific enzymatic hydrolysis of legume proteins, resulting in the production of hydrolysates with diverse functional properties, including solubility, emulsibility, gelability, and foamability, which are determined by the composition (peptide/amino acid) and structure (secondary/tertiary) of legume proteins after LAB fermentation. The correlation between LAB-specific enzymatic hydrolysis, protein composition and structure, and protein functional properties will assist in selecting legume protein raw materials and LAB strains for legume plant-based food products and expand the application of legume proteins in the food industry.

CLI: A new protocol for the isolation of Lactic Acid Bacteria from complex plant samples.

Lactic Acid Bacteria (LAB) are predominantly probiotic microorganisms and the most are Generally Recognized As Safe (GRAS). LAB inhabit in the human gut ecosystem and are largely found in fermented foods and silage. In the last decades, LAB have also has been found in plant microbiota as a new class of microbes with probiotic activity to plants. For this reason, today the scientific interest in the study and isolation of LAB for agronomic application has increased. However, isolation protocols from complex samples such as plant tissues are scarce and inefficient. In this study, we developed a new protocol (CLI, Complex samples LAB Isolation) which yields purified LAB from plants. The sensitivity of CLI protocol was sufficient to isolate representative microorganisms of LAB genera (i.e. Leuconostoc, Lactococcus and Enterococcus). CLI protocol consists on five steps: i) sample preparation and pre-incubation in 1% sterile peptone at 30 °C for 24-48 h; ii) Sample homogenization in vortex by 10 min; iii) sample serial dilution in quarter-strength Ringer solution, iv) incubation in MRS agar plates with 0.2% of sorbic acid, with 1% of CaCO3, O2 < 15%, at pH 5.8 and 37 °C for 48 h.; v) Selection of single colonies with LAB morphology and CaCO3-solubilization halo. Our scientific contribution is that CLI protocol could be used for several complex samples and represents a useful method for further studies involving native LAB.

Lactic acid bacteria reduce bacterial diarrhea in rabbits via enhancing immune function and restoring intestinal microbiota homeostasis.

BACKGROUND: Numerous previous reports have demonstrated the efficacy of Lactic acid bacteria (LAB) in promoting growth and preventing disease in animals. In this study, Enterococcus faecium ZJUIDS-R1 and Ligilactobaciiius animalis ZJUIDS-R2 were isolated from the feces of healthy rabbits, and both strains showed good probiotic properties in vitro. Two strains (108CFU/ml/kg/day) were fed to weaned rabbits for 21 days, after which specific bacterial infection was induced to investigate the effects of the strains on bacterial diarrhea in the rabbits. RESULTS: Our data showed that Enterococcus faecium ZJUIDS-R1 and Ligilactobaciiius animalis ZJUIDS-R2 interventions reduced the incidence of diarrhea and systemic inflammatory response, alleviated intestinal damage and increased antibody levels in animals. In addition, Enterococcus faecium ZJUIDS-R1 restored the flora abundance of Ruminococcaceae1. Ligilactobaciiius animalis ZJUIDS-R2 up-regulated the flora abundance of Adlercreutzia and Candidatus Saccharimonas. Both down-regulated the flora abundance of Shuttleworthia and Barnesiella to restore intestinal flora balance, thereby increasing intestinal short-chain fatty acid content. CONCLUSIONS: These findings suggest that Enterococcus faecium ZJUIDS-R1 and Ligilactobaciiius animalis ZJUIDS-R2 were able to improve intestinal immunity, produce organic acids and regulate the balance of intestinal flora to enhance disease resistance and alleviate diarrhea-related diseases in weanling rabbits.

Investigating human-derived lactic acid bacteria for alcohol resistance.

BACKGROUND: Excessive alcohol consumption has been consistently linked to serious adverse health effects, particularly affecting the liver. One natural defense against the detrimental impacts of alcohol is provided by alcohol dehydrogenase (ADH) and acetaldehyde dehydrogenase (ALDH), which detoxify harmful alcohol metabolites. Recent studies have shown that certain probiotic strains, notably Lactobacillus spp., possess alcohol resistance and can produce these critical enzymes. Incorporating these probiotics into alcoholic beverages represents a pioneering approach that can potentially mitigate the negative health effects of alcohol while meeting evolving consumer preferences for functional and health-centric products. RESULTS: Five lactic acid bacteria (LAB) isolates were identified: Lactobacillus paracasei Alc1, Lacticaseibacillus rhamnosus AA, Pediococcus acidilactici Alc3, Lactobacillus paracasei Alc4, and Pediococcus acidilactici Alc5. Assessment of their alcohol tolerance, safety, adhesion ability, and immunomodulatory effects identified L. rhamnosus AA as the most promising alcohol-tolerant probiotic strain. This strain also showed high production of ADH and ALDH. Whole genome sequencing analysis revealed that the L. rhamnosus AA genome contained both the adh (encoding for ADH) and the adhE (encoding for ALDH) genes. CONCLUSIONS: L. rhamnosus AA, a novel probiotic candidate, showed notable alcohol resistance and the capability to produce enzymes essential for alcohol metabolism. This strain is a highly promising candidate for integration into commercial alcoholic beverages upon completion of comprehensive safety and functionality evaluations.

Reducing transmission of high-risk antibiotic resistance genes in whole-crop corn silage through lactic acid bacteria inoculation and increasing ensiling temperature.

The microbial hosts of antibiotic resistance genes (ARGs) found epiphytically on plant materials could grow and flourish during silage fermentation. This study employed metagenomic analysis and elucidated the occurrence and transmission mechanisms of ARGs and their microbial hosts in whole-crop corn silage inoculated with homofermentative strain Lactiplantibacillus plantarum or heterofermentative strain Lentilactobacillus buchneri ensiled under different temperature (20 and 30 °C). The results revealed that the corn silage was dominated by Lactobacillus, Leuconostoc, Lentilactobacillus, and Latilactobacillus. Both the ensiling temperature and inoculation had greatly modified the silage microbiota. However, regardless of the ensiling temperature, L. buchneri had significantly higher ARGs, while it only exhibited significantly higher mobile genetic elements (MGEs) in low temperature treatments. The microbial community of the corn silage hosted highly diverse form of ARGs, which were primarily MacB, RanA, bcrA, msbA, TetA (58), and TetT and mainly corresponded to macrolides and tetracyclines drug classes. Plasmids were identified as the most abundant MGEs with significant correlation with some high-risk ARGs (tetM, TolC, mdtH, and NorA), and their abundances have been reduced by ensiling process. Furthermore, higher temperature and L. buchneri reduced abundances of high-risk ARGs by modifying their hosts and reduced their transmission in the silage. Therefore, ensiling, L. buchneri inoculation and higher storage temperature could improve the biosafety of corn silage.

Lactic acid bacteria modulate the CncC pathway to enhance resistance to ß-cypermethrin in the oriental fruit fly.

The gut microbiota of insects has been shown to regulate host detoxification enzymes. However, the potential regulatory mechanisms involved remain unknown. Here, we report that gut bacteria increase insecticide resistance by activating the cap "n" collar isoform-C (CncC) pathway through enzymatically generated reactive oxygen species (ROS) in Bactrocera dorsalis. We demonstrated that Enterococcus casseliflavus and Lactococcus lactis, two lactic acid-producing bacteria, increase the resistance of B. dorsalis to ß-cypermethrin by regulating cytochrome P450 (P450) enzymes and α-glutathione S-transferase (GST) activities. These gut symbionts also induced the expression of CncC and muscle aponeurosis fibromatosis. BdCncC knockdown led to a decrease in resistance caused by gut bacteria. Ingestion of the ROS scavenger vitamin C in resistant strain affected the expression of BdCncC/BdKeap1/BdMafK, resulting in reduced P450 and GST activity. Furthermore, feeding with E. casseliflavus or L. lactis showed that BdNOX5 increased ROS production, and BdNOX5 knockdown affected the expression of the BdCncC/BdMafK pathway and detoxification genes. Moreover, lactic acid feeding activated the ROS-associated regulation of P450 and GST activity. Collectively, our findings indicate that symbiotic gut bacteria modulate intestinal detoxification pathways by affecting physiological biochemistry, thus providing new insights into the involvement of insect gut microbes in the development of insecticide resistance.

Reducing Immunoreactivity of Gluten Peptides by Probiotic Lactic Acid Bacteria for Dietary Management of Gluten-Related Diseases.

Immunoreactive gluten peptides that are not digested by peptidases produced by humans can trigger celiac disease, allergy and non-celiac gluten hypersensitivity. The aim of this study was to evaluate the ability of selected probiotic strains to hydrolyze immunoreactive gliadin peptides and to identify peptidase-encoding genes in the genomes of the most efficient strains. Residual gliadin immunoreactivity was measured after one- or two-step hydrolysis using commercial enzymes and bacterial peptidase preparations by G12 and R5 immunoenzymatic assays. Peptidase preparations from Lacticaseibacillus casei LC130, Lacticaseibacillus paracasei LPC100 and Streptococcus thermophilus ST250 strains significantly reduced the immunoreactivity of gliadin peptides, including 33-mer, and this effect was markedly higher when a mixture of these strains was used. In silico genome analyses of L. casei LC130 and L. paracasei LPC100 revealed the presence of genes encoding peptidases with the potential to hydrolyze bonds in proline-rich peptides. This suggests that L. casei LC130, L. paracasei LPC100 and S. thermophilus ST250, especially when used as a mixture, have the ability to hydrolyze immunoreactive gliadin peptides and could be administered to patients on a restricted gluten-free diet to help treat gluten-related diseases.

Brazilian indigenous nonstarter lactic acid bacteria enhance the diversification of volatile compounds in short-aged cheese.

There is growing interest in using autochthonous lactic acid bacteria (LAB) that provide unique sensory characteristics to dairy products without affecting their safety and quality. This work studied the capacity of three Brazilian indigenous nonstarter LABs (NSLAB) to produce biogenic amines (BAs) and evaluated their effect on the volatile organic compounds (VOCs), microbial LAB communities, and physicochemical profile of short-aged cheese. Initially, the strain's potential for biosynthesis of BAs was assessed by PCR and in vitro assays. Then, a pilot-scale cheese was produced, including the NSLAB, and the microbial and VOC profiles were analyzed after 25 and 45 days of ripening. As a results, the strains did not present genes related to relevant BAs and did not produce them in vitro. During cheese ripening, the Lactococci counts were reduced, probably in the production of alcohols and acid compounds by the NSLAB. Each strain produces a unique VOC profile that changes over the ripening time without the main VOCs related to rancid or old cheese. Particularly, the use of the strain Lacticaseibacillus. paracasei ItalPN16 resulted in production of ester compounds with fruity notes. Thus, indigenous NSLAB could be a valuable tool for the enhancement and diversification of flavor in short-aged cheese.

Impact of lactic acid bacteria inoculation on fungal diversity during Spanish-style green table olive fermentations.

This study delved into the evolution of fungal population during the fermentation of Spanish-style green table olives (Manzanilla cultivar), determining the influence of different factors such as fermentation matrix (brine or fruit) or the use of a lactic acid bacteria inoculum, on its distribution. The samples (n = 24) were directly obtained from industrial fermentation vessels with approximately 10.000 kg of fruits and 6.000 L of brines. Our findings showcased a synchronized uptick in lactic acid bacteria counts alongside fungi proliferation. Metataxonomic analysis of the Internal Transcribed Spacer (ITS) region unearthed noteworthy disparities across different fermentation time points (0, 24, and 83 days). Statistical analysis pinpointed two Amplicon Sequence Variants (ASV), Candida and Aureobasidium, as accountable for the observed variances among the different fermentation time samples. Notably, Candida exhibited a marked increase during 83 days of fermentation, opposite to Aureobasidium, which demonstrated a decline. Fungal biodiversity was slightly higher in brines than in fruits, whilst no effect of inoculation was noticed. At the onset of fermentation, prominently detected genera were also Mycosphaerella (19.82 %) and Apohysomyces (16.31 %), hitherto unreported in the context of table olive processing. However, their prevalence dwindled to nearly negligible levels from 24th day fermentation onwards (<2 %). On the contrary, they were replaced by the fermentative yeasts Saccharomyces and Isstachenkia. Results obtained in this work will be useful for designing new strategies for better control of table olive fermentations.

Cereus jamacaru DC. (mandacaru) fruit as a source of lactic acid bacteria with in vitro probiotic-related characteristics and its protective effects on Pediococcus pentosaceus during lyophilization and refrigeration storage.

This study isolated and identified autochthonous lactic acid bacteria (LAB) from mandacaru fruit and evaluated their potential probiotic and technological aptitudes in vitro, as well as the protective effects of freeze-dried mandacaru fruit on the most promising LAB isolate during lyophilization and refrigeration storage. Initially, 212 colonies were isolated from mandacaru fruit, and 34 were preliminarily identified as LAB. Thirteen isolates identified by 16S-rRNA sequencing as Pediococcus pentosaceus were negative for DNase, gelatinase, hemolytic, and biogenic amine production. The selected isolates showed proteolytic activity, diacetyl and exopolysaccharide production, and good tolerance to different NaCl concentrations while having low cellular hydrophobicity and antagonistic activity against pathogens. The survival of isolates sharply decreased after 3 h of exposure to pH 2 and had a good tolerance to 1 % bile salt. A principal component analysis selected P. pentosaceus 57 as the most promising isolate based on the examined technological and probiotic-related physiological properties. This isolate was lyophilized with mandacaru fruit and stored under refrigeration for 90 days. P. pentosaceus 57 lyophilized with mandacaru fruit had high viable cell counts (9.69 ± 0.03 log CFU/mL) and >50 % of physiologically active cells at 90 days of refrigeration storage. The results indicate that mandacaru fruit is a source of P. pentosaceus with aptitudes to be explored as potential probiotic and technological characteristics of interest for the food industry, besides being a good candidate for use in lyophilization processes and refrigeration storage of LAB due to its cryoprotective effects.

Study and characterization of a product based on a vegetable extract of quinoa fermented with water kefir grains.

This work aimed to study and characterize a product based on vegetable extract of quinoa (WVEQ) fermented with water kefir grains. The effect of sucrose concentration (SC), inulin concentration (IC), and xanthan gum (XG) concentration were evaluated using a central composite design (CCD) 23. They were subsequently characterized regarding cellular growth of the grains, beverage yield, pH, soluble solids, carbon dioxide (CO2) production, lactic acid, and ethanol production. Therefore, for the final stage, two formulations (F1 and F8) of the CCD were chosen to be characterized in terms of proximate composition, microbiological composition of the kefir culture, analysis of organic compounds, sensory analysis, and enzymatic and microbiological characterization before and after simulation of in vitro gastrointestinal digestion. In the two chosen products, one can see that fermentation optimized the bioavailability of proteins due to the high proteolytic activity of the microorganisms in kefir and the increase in lipid content. In identifying microorganisms, there was a prevalence of Saccharomyces sp. yeasts. In the sensory analysis, the F8 formulation showed better results than the F1 formulation. In vitro, gastrointestinal digestion showed reduced lactic acid bacteria and yeast and increased acetic acid bacteria in the liquid phase for both formulations. In the enzymatic profile, there was a reduction in all enzymes analyzed for both formulations, except for amylase in F1, which went from 14.05 U/mL to 39.41 U/mL. Therefore, it is concluded that using WVEQ as a substrate for the product appears to be a viable alternative with nutritional and technological advantages for serving a specific market niche.