Search Results (19)

Campylobacter spp. are prevalent foodborne bacterial enteric pathogens. Their inclusion in biofilms on abiotic surfaces is considered a strategy that facilitates their extraintestinal survival. Organic acid (OA) treatments could be used in a green approach to decontaminate various surfaces. This work aimed to evaluate the inhibitory and eradicative effects of L(+)-lactic acid (LA), a naturally occurring OA, on a dual-species biofilm formed on two food processing model surfaces (polystyrene and stainless steel) by three selected foodborne Campylobacter spp. isolates (two C. jejuni and one C. coli). The influence of aerobiosis conditions (microaerophilic, aerobic and CO₂ enriched) on the resistance of the established biofilms to the acid was also tested. In parallel, the predominant metabolites contained in the planktonic media of biofilm monocultures and mixed-culture biofilm were comparatively analyzed by an untargeted metabolomics approach. Results revealed that LA inhibited mixed-culture biofilm formation by more than 2 logs (>99%) on both surfaces when this was applied at its highest tested concentration (4096 μg/mL; 0.34% v/v). However, all the preformed mixed-culture biofilms (ca. 10⁶⁻⁷ CFU/cm²) could not be eradicated even when the acid was used at concentrations exceeding 5% v/v, denoting their extremely high recalcitrance which was still influenced by the abiotic substratum, and the biofilm-forming aerobiosis conditions. The metabolic analysis revealed a strain-specific metabolite production which might also be related to the strain-specific biofilm-forming and resistance behaviors and resulted in the distinct clustering of the different samples. Overall, the current findings provide important information on the effectiveness of LA against biofilm campylobacteria and may assist in mitigating their risk in the food chain. Full article

Acetoin biosynthesis by two Bacillus subtilis strains valorising crude glycerol was thoroughly explored within a pre-defined range of culture conditions and systems. B. subtilis ACA-DC 1176 stood out for its higher efficiency in acetoin production, prompting an investigation into the potential for enhanced productivity through the evaluation of diverse culture conditions and media compositions. The primary by-products of the biodiesel and corn industries, namely crude glycerol and corn steep liquor, respectively, were successfully employed as the principal carbon and nitrogen sources of the newly developed low-cost culture medium. Furthermore, the results of the various feeding strategies that were tested indicated that the conversion of 2,3-butanediol (BDO) to acetoin occurred exclusively when the concentration of glycerol was below approximately 5 g/L. This seemed to be necessary for the production of NADH, which is essential for maintaining cellular processes. Following the complete depletion of glycerol, acetic acid increased and became the predominant metabolite, while both acetoin and BDO decreased, presumably resulting in ATP generation. This is likely a mechanism employed by the cell to generate energy in the absence of a carbon source. In the fed-batch bioreactor culture, the kinetics of metabolites differed, as there was no conversion of BDO to acetoin at the final depletion of glycerol. At volumetric mass transfer coefficient (k_La) levels exceeding approximately 70 1/h, the production of acetoin was favoured over that of BDO, with the highest observed acetoin/BDO ratio reaching 4.29 g/g. Conversely, at k_La values below approximately 60 1/h, the titres of acetoin and BDO were found to be nearly equal. The final concentrations of acetoin and BDO reached 36.0 g/L and 25.5 g/L, respectively, resulting in a total yield of both (acetoin + BDO) per glycerol consumption of 0.40 g/g. To the best of our knowledge, this is the first study to focus on acetoin production from crude glycerol fermentative valorisation. The study presents new findings regarding the parameters influencing the level of BDO conversion to acetoin. However, further research is required in order to gain a comprehensive understanding of the underlying phenomena and metabolic pathways involved. Full article

Pseudomonas aeruginosa lung infections in cystic fibrosis (CF) patients represent a therapeutic challenge due to antibiotic resistance. Repurposing existing drugs is a promising approach for identifying new antimicrobials. A crucial factor in successful drug repurposing is using assay conditions that mirror the site of infection. Here, the impact of growth conditions on the anti-P. aeruginosa activity of a library of 3386 compounds was evaluated. To this, after 24 h exposure, the survival rate of CF P. aeruginosa RP73 planktonic cells was assessed spectrophotometrically under “CF-like” (artificial CF sputum, pH 6.8, 5% CO₂) and enriched (Tryptone Soya Broth, pH 7.2, and aerobiosis) conditions. Among non-antibiotic compounds (n = 3127), 13.4% were active regardless of growth conditions, although only 3.2% had comparable activity; 4% and 6.2% were more active under CF-like or enriched conditions, respectively. Interestingly, 22.1% and 26.6% were active exclusively under CF-like and enriched conditions, respectively. Notably, 7 and 12 hits caused 100% killing under CF-like and enriched conditions, respectively. Among antibiotics (n = 234), 42.3% were active under both conditions, although only 18.4% showed comparable activity; 9.4% and 14.5% were more active under CF-like and enriched conditions, respectively. Interestingly, 23% and 16.6% were active exclusively under CF-like and enriched conditions, respectively. Sulphonamides showed higher activity under CF-like conditions, whereas tetracyclines, fluoroquinolones, and macrolides were more effective under enriched settings. Our findings indicated that growth conditions significantly affect the anti-P. aeruginosa activity of antibiotics and non-antibiotic drugs. Consequently, repurposing studies and susceptibility tests should be performed under physicochemical conditions that the pathogen tackles at the site of infection. Full article

The primary goal of this scoping review is to collect, analyze, and critically describe information regarding the role of the main compounds (reuterin, phenyllactic acid, and exopolysaccharides) produced by LAB that possess antifungal properties and provide some suggestions for further research. The use of lactic acid bacteria (LAB) to mitigate spoilage and extend the shelf life of foodstuffs has a long history. Recently, there has been a growing interest in the unique properties of these additions to the foodstuffs in which they are applied. In recent studies regarding biopreservation, significant attention has been given to the role of these microorganisms and their metabolites. This fascinating recent discipline aims not only to replace traditional preservation systems, but also to improve the overall quality of the final product. The biologically active by-products produced by lactic acid bacteria are synthesized under certain conditions (time, temperature, aerobiosis, acidity, water activity, etc.), which can be enacted through one of the oldest approaches to food processing: fermentation (commonly used in the dairy and bakery sectors). This study also delves into the biosynthetic pathways through which they are synthesized, with a particular emphasis on what is known about the mechanisms of action against molds in relation to the type of food. Full article

The aim of this study was to assess the antimicrobial effects of myrtle (Myrtus communis L.) essential oil (EO) on pathogenic (E. coli O157:H7 NCTC 12900; Listeria monocytogenes ATCC BAA-679) and spoilage microbiota in beef and determine its minimum inhibitory concentration (MIC) and antioxidant activity. The behavior of LAB, Enterobacteriaceae, Pseudomonas spp., and fungi, as well as total mesophilic (TM) and total psychotropic (TP) counts, in beef samples, was analyzed during storage at 2 and 8 °C in two different packaging systems (aerobiosis and vacuum). Leaves of myrtle were dried, its EO was extracted by hydrodistillation using a Clevenger-type apparatus, and the chemical composition was determined using chromatographical techniques. The major compounds obtained were myrtenyl acetate (15.5%), β-linalool (12.3%), 1,8-cineole (eucalyptol; 9.9%), geranyl acetate (7.4%), limonene (6.2%), α-pinene (4.4%), linalyl o-aminobenzoate (5.8%), α-terpineol (2.7%), and myrtenol (1.2%). Myrtle EO presented a MIC of 25 µL/mL for E. coli O157:H7 NCTC 12900, E. coli, Listeria monocytogenes ATCC BAA-679, Enterobacteriaceae, and E. coli O157:H7 ATCC 35150 and 50µL/mL for Pseudomonas spp. The samples packed in aerobiosis had higher counts of deteriorative microorganisms than samples packed under vacuum, and samples with myrtle EO presented the lowest microbial contents, indicating good antimicrobial activity in beef samples. Myrtle EO is a viable natural alternative to eliminate or reduce the pathogenic and deteriorative microorganisms of meat, preventing their growth and enhancing meat safety. Full article

Cytosolic pyruvate is an essential metabolite in lactic acid production during microbial fermentation. However, under aerobiosis, pyruvate is transported to the mitochondrial matrix by the mitochondrial pyruvate carrier (MPC) and oxidized in cell respiration. Previous reports using Saccharomyces cerevisiae or Aspergillus oryzae have shown that the production of pyruvate-derived chemicals is improved by deleting the MPC1 gene. A previous lactate-producing K. phaffii strain engineered by our group was used as a host for the deletion of the MPC1 gene. In addition, the expression of a bacterial hemoglobin gene under the alcohol dehydrogenase 2 promoter from Scheffersomyces stipitis, known to work as a hypoxia sensor, was used to evaluate whether aeration would supply enough oxygen to meet the metabolic needs during lactic acid production. However, unlike S. cerevisiae and A. oryzae, the deletion of Mpc1 had no significant impact on lactic acid production but negatively affected cell growth in K. phaffii strains. Furthermore, the relative quantification of the VHb gene revealed that the expression of hemoglobin was detected even in aerobic cultivation, which indicates that the demand for oxygen in the bioreactor could result in functional hypoxia. Overall, the results add to our previously published ones and show that blocking cell respiration using hypoxia is more suitable than deleting Mpc for producing lactic acid in K. phaffii. Full article

γ-aminobutyric acid (GABA) has several beneficial effects on human health. GABA may be produced via chemical synthesis or through microbial metabolism, and Levilactobacillus brevis is recognized as a GABA-producing species. In this study, 11 Lvb. brevis strains were screened for GABA production, and the best producers were selected to verify the effect of aerobic (AE) and respiratory (RS) cultivations on growth parameters, biomass, and GABA accumulation. Lvb. brevis LB12 was then used to evaluate the combined effect of the incubation atmosphere (anaerobiosis vs. aerobiosis), cell protection (free vs. immobilized cells), and cell recycling (fresh vs. starved cells) on GABA production. Glutamate (GLU) consumption and GABA accumulation were detected by Thin-layer Chromatography (TLC) and RP-HPLC analyses. The ability to produce GABA was widespread among the strains. AE and RS growth improved biomass production, but oxygen availability impaired GLU to GABA conversion, and the anaerobically growing cells had the highest GABA productivity. Immobilized strains had lower efficiency in both GLU uptake and conversion compared to free cells, probably due to the poor diffusion in alginate beads. The use of resting cells allowed further GABA production without the cultivation step, but cell activity was exhausted after three cycles of reutilization. Lvb. brevis LB12 is an excellent GABA producer, and AE cultivation can be exploited to improve the final cell density; however, the conditions for boosting GLU to GABA conversion and cell regeneration need to be further investigated. Full article

The chemical composition of cactus forage becomes a favorable culture medium for accelerated microbial activity when exposed to air, as it contains high content of non-fiber carbohydrates and water. Thus, the aim of this study was to evaluate the bacterial community dynamics of different mixtures, using fresh forage of cactus and buffel grass hay as a function of the period of exposure to air. The experimental design used was a 5 × 5 factorial completely randomized (five levels of cactus forage × five times of exposure to air), with five replications. The peak of Escherichia coli population growth was after 16.06 h of exposure to air, observed in treatments of 90% and 100% cactus forage. There was an increase in microbial richness and uniformity of all treatments after six hours. The most abundant genera were Weissella, Lactobacillus, Bacteroides, Pseudomonas, Sphingobacterium, and Sphingomonas. The diet with 100% cactus forage showed a predominance of Weissella, Lactobacillus, and Leuconostoc. With 20% cactus forage, there was a greater apparent abundance of Pseudomonas, Sphingomonas, and Sphingobacterium. Aerobic exposure of mixtures of cactus forage with buffel grass hay increases the proliferation of microorganisms with pathogenic potential in the diet. Aerobic exposure of mixtures of cactus forage with buffel grass hay increases the proliferation of microorganisms with pathogenic potential in the diet. Therefore, an exposure period of fewer than six hours with 20% cactus forage is recommended to minimize levels of E. coli. Avoiding negative effects of the multiplication of pathogenic microorganisms on animal and human health. Full article

The dyeing processes of the textile industry generate waste products such as unfixed dyes, phenolic surfactants and heavy metals. These constitute an environmental problem for the bodies receiving their wastewater due to the interruption of the lighting in the aquatic environment and the release of toxic molecules by the decomposition of the dyes. There are several treatment methods, of which biological methods are the most feasible. In the current study, the I5-ESPE microbial consortium was obtained and evaluated on the components of textile wastewater, in addition to the selection of a support for an anaerobic reactor that is directed to the treatment of effluents from the textile industry. Two microbial consortia were achieved by exposure to air in Pseudomonas culture medium modified with direct dyes Red 23 and Blue 106, evaluating their removal capacity of the reactive dyes Navy 171, Red 141 and Yellow 84. The consortium I5-ESPE was selected for its greatest action, yielding approximately 95% removal. Its tolerance to phenol was also determined; we reached 98% removal of chromium(VI) and 67% of total chromium under anaerobic conditions and some 25% zinc in aerobiosis. The reduction in the chemical oxygen demand (COD) was evaluated with (57.03%) and without (31.47%) aeration. The species Staphylococcus xylosus, Saccharomyces cerevisiae and Candida tropicalis were identified prior to treatment of textile wastewater, as well as Enterobacter cloacae and Bacillus megaterium after treatment. Bacillus subtilis was present throughout the process. We evaluated coconut shell as a support for an anaerobic reactor, and it demonstrated better physical characteristics than plastic and common rock, in addition to similar results in the reduction in COD of 50%, volatile suspended solids of 2545.46 mg/L and total suspended solids of 282.82 mg/L. Full article

Infections caused by Mycobacterium abscessus (Mab), an environmental non-tuberculous mycobacterium, are difficult to eradicate from patients with pulmonary diseases such as cystic fibrosis and bronchiectasis even after years of antibiotic treatments. In these people, the low oxygen pressure in mucus and biofilm may restrict Mab growth from actively replicating aerobic (A) to non-replicating hypoxic (H) stages, which are known to be extremely drug-tolerant. After the exposure of Mab A and H cells to drugs, killing was monitored by measuring colony-forming units (CFU) and regrowth in liquid medium (MGIT 960) of 1-day-old A cells (A1) and 5-day-old H cells (H5). Mab killing was defined as a lack of regrowth of drug-exposed cells in MGIT tubes after >50 days of incubation. Out of 18 drugs tested, 14-day treatments with bedaquiline-amikacin (BDQ-AMK)-containing three-drug combinations were very active against A1 + H5 cells. However, drug-tolerant cells (persisters) were not killed, as shown by CFU curves with typical bimodal trends. Instead, 56-day treatments with the nitrocompounds containing combinations BDQ-AMK-rifabutin-clarithromycin-nimorazole and BDQ-AMK-rifabutin-clarithromycin-metronidazole-colistin killed all A1 + H5 Mab cells in 42 and 56 days, respectively, as shown by lack of regrowth in agar and MGIT medium. Overall, these data indicated that Mab persisters may be killed by appropriate drug combinations. Full article

Leuconostoc mesenteroides includes strains used as starter and/or adjunct cultures for the production of several fermented foods. In this study, the effect of anaerobic and respiratory cultivations, as well as of citrate supplementation and different pH values, was evaluated on growth, biomass, metabolite, and enzymatic activities (pyruvate oxidase, POX; NADH-dependent oxidase, NOX; NADH-dependent peroxidase, NPR) of Leuconostoc mesenteroides subsp. cremoris E30. We compared the respiration-increased growth rate and biomass production of Leuc. mesenteroides E30 to anaerobic cultivation. A supplementation of citrate impaired the growth rate of the respiratory cells. As expected, anaerobic cultures did not consume oxygen, and a similar trend in oxygen uptake was observed in respiratory cultures. The aerobic incubation caused changes in the metabolic pattern, reducing the production of ethanol in favour of acetic acid. Citrate was already exhausted in the exponential phase and did not affect the yields in acetic acid and ethanol. NOX activity increased in the presence of oxygen, while catalase was also detected in the absence of hemin. The absence of H₂O₂ suggested its degradation by NPR and catalase. Respiratory cultivation provided benefits (increase in growth rate, biomass, and activity in antioxidant enzymes) for Leuc. mesenteroides E30. Therefore, the exploitation of respiratory phenotypes may be useful for the formulation of competitive starter or adjunct cultures. Full article

Arc (anoxic redox control), one of the most intensely investigated two-component regulatory systems in γ-proteobacteria, plays a major role in mediating the metabolic transition from aerobiosis to anaerobiosis. In Shewanella oneidensis, a research model for respiratory versatility, Arc is crucial for aerobic growth. However, how this occurs remains largely unknown. In this study, we demonstrated that the loss of the response regulator ArcA distorts the correlation between transcription and translation by inhibiting the ribosome biosynthesis. This effect largely underlies the growth defect because it concurs with the effect of chloramphenicol, which impairs translation. Reduced transcription of ArcA-dependent ribosomal protein S1 appears to have a significant impact on ribosome assembly. We further show that the lowered translation efficiency is not accountable for the envelope defect, another major defect resulting from the ArcA loss. Overall, our results suggest that although the arcA mutation impairs growth through multi-fold complex impacts in physiology, the reduced translation efficacy appears to be a major cause for the phenotype, demonstrating that Arc is a primary system that coordinates proteomic resources with metabolism in S. oneidensis. Full article

The biotechnological production of dicarboxylic acids (C4) from renewable carbon sources represents an attractive approach for the provision of these valuable compounds by green chemistry means. Glycerol has become a waste product of the biodiesel industry that serves as a highly reduced carbon source for some microorganisms. Escherichia coli is capable of consuming glycerol to produce succinate under anaerobic fermentation, but with the deletion of some tricarboxylic acid (TCA) cycle genes, it is also able to produce succinate and malate in aerobiosis. In this study, we investigate possible rate-limiting enzymes by overexpressing the C-feeding anaplerotic enzymes Ppc, MaeA, MaeB, and Pck in a mutant that lacks the succinate dehydrogenase (Sdh) enzyme. The overexpression of the TCA enzyme Mdh and the activation of the glyoxylate shunt was also examined. Using this unbiased approach, we found that phosphoenol pyruvate carboxylase (Ppc) overexpression enhances an oxidative pathway that leads to increasing succinate, while phosphoenol pyruvate carboxykinase (Pck) favors a more efficient reductive branch that produces mainly malate, at 57.5% of the theoretical maximum molar yield. The optimization of the culture medium revealed the importance of bicarbonate and pH in the production of malate. An additional mutation of the ppc gene highlights its central role in growth and C4 production. Full article

The ability of Pseudomonas aeruginosa to form biofilm during a long-term infection makes it difficult to treat patients correctly. The current clinical antimicrobial susceptibility testing methods are based on the study of planktonic strains. A standardized protocol to analyze the antimicrobial susceptibility in biofilms is necessary for routine laboratories. The aims of this study were to develop a simple biofilm model and to study the antimicrobial susceptibility of P. aeruginosa strains in biofilm growth. Different artificial sputum media, and aerobiosis and microaerobiosis conditions were analyzed using a microtiter plate method and P. aeruginosa PAO1 as reference strain. Planktonic and biofilm antimicrobial susceptibility to cefepime, imipenem, azithromycin, gentamicin, tobramycin, and ciprofloxacin were determined in clinical and non-clinical P. aeruginosa strains. The Synthetic Cystic Fibrosis Medium was proposed as a good medium. The biofilm greatly increased the resistance to tested antimicrobials, except for azithromycin. Cefepime and imipenem showed poor anti-biofilm effect while tobramycin, gentamicin, and ciprofloxacin showed good activity in some strains. Azithromycin showed a better activity in biofilm than in planktonic state when aerobic conditions were used. This study establishes useful information to test antimicrobial susceptibility in P. aeruginosa biofilms, and includes possible antimicrobial options to treat long-term infected patients. Full article

A triathlon is an extremely high-intensity exercise and a challenge for physiological adaptation. A triathlete’s microbiome might be modulated by diet, age, medical treatments, lifestyle, and exercise, thereby maintaining aerobiosis and optimum health and performance. Probiotics, prebiotics, and synbiotics have been reported to have health-promoting activities (e.g., immunoregulation and cancer prevention). However, few studies have addressed how probiotics affect the microbiota of athletes and how this translates into functional activities. In our previous study, we found that Lactobacillus plantarum PS128 could ameliorate inflammation and oxidative stress, with improved exercise performance. Thus, here we investigate how the microbiota of triathletes are altered by L. plantarum PS128 supplementation, not only for exercise performance but also for possible physiological adaptation. The triathletes were assigned to two groups: an L. plantarum 128 supplement group (LG, 3 × 10¹⁰ colony-forming units (CFU)/day) and a placebo group (PG). Both groups continued with their regular exercise training for the next 4 weeks. The endurance performance, body composition, biochemistries, blood cells, microbiota, and associated metabolites were further investigated. PS128 significantly increased the athletes’ endurance, by about 130% as compared to the PG group, but there was no significant difference in maximal oxygen consumption (VO₂max) and composition between groups. The PS128 supplementation (LG) modulated the athlete’s microbiota with both significant decreases (Anaerotruncus, Caproiciproducens, Coprobacillus, Desulfovibrio, Dielma, Family_XIII, Holdemania, and Oxalobacter) and increases (Akkermansia, Bifidobacterium, Butyricimonas, and Lactobacillus), and the LG showed lower diversity when compared to the PG. Also, the short-chain fatty acids (SCFAs; acetate, propionate, and butyrate) of the LG were significantly higher than the PG, which might be a result of a modulation of the associated microbiota. In conclusion, PS128 supplementation was associated with an improvement on endurance running performance through microbiota modulation and related metabolites, but not in maximal oxygen uptake. Full article