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Adaptation and tolerance of bacteria against acetic acid
Abstract
Acetic acid is a weak organic acid exerting a toxic effect to most microorganisms at concentrations as low as 0.5 wt%. This toxic effect results mostly from acetic acid dissociation inside microbial cells, causing a decrease of intracellular pH and metabolic disturbance by the anion, among other deleterious effects. These microbial inhibition mechanisms enable acetic acid to be used as a preservative, although its usefulness is limited by the emergence of highly tolerant spoilage strains. Several biotechnological processes are also inhibited by the accumulation of acetic acid in the growth medium including production of bioethanol from lignocellulosics, wine making, and microbe-based production of acetic acid itself. To design better preservation strategies based on acetic acid and to improve the robustness of industrial biotechnological processes limited by this acid's toxicity, it is essential to deepen the understanding of the underlying toxicity mechanisms. In this sense, adaptive responses that improve tolerance to acetic acid have been well studied in Escherichia coli and Saccharomyces cerevisiae. Strains highly tolerant to acetic acid, either isolated from natural environments or specifically engineered for this effect, represent a unique reservoir of information that could increase our understanding of acetic acid tolerance and contribute to the design of additional tolerance mechanisms. In this article, the mechanisms underlying the acetic acid tolerance exhibited by several bacterial strains are reviewed, with emphasis on the knowledge gathered in acetic acid bacteria and E. coli. A comparison of how these bacterial adaptive responses to acetic acid stress fit to those described in the yeast Saccharomyces cerevisiae is also performed. A systematic comparison of the similarities and dissimilarities of the ways by which different microbial systems surpass the deleterious effects of acetic acid toxicity has not been performed so far, although such exchange of knowledge can open the door to the design of novel approaches aiming the development of acetic acid-tolerant strains with increased industrial robustness in a synthetic biology perspective.
... According to the Daqu fermentation process conditions, the Daqu samples at different fermentation periods (1,2,4,6,7,8,10,13,17,21, and 25 days) were collected from the Daqu room of Baijiu winery in YiBin. The samples were crushed and mixed into sterile sampling bags and stored at -80°C. ...
... On the other hand, Candida regulates ethyl lactate production in Daqu(X., while Acetobacter and Gluconobacter oxidize ethanol to acetic acid(Trcek, Mira, & Jarboe, 2015) and contribute to the formation of ethyl acetate, the main avor substance in Baijiu. Thus, LAB contributes to the shaping and succession of microbial community structure during Daqu fermentation and in uences its avor.3.4. ...
OBJECTIVES: The microbial community structure of the saccharifying starter, Nongxiangxing Daqu（Daqu）, is a crucial factor that determines the quality of Baijiu. lactic acid bacteria (LAB), the dominant microorganisms in the Daqu. The present study investigated the effects of LAB on the microbial community structure and its contribution to microbial community function during the fermentation of Daqu.
RESULTS: Box-line variance analysis shows that LAB showed a significant stage-specific succession pattern during Daqu fermentation. The LEfSe analysis and the random forest learning algorithm identified LAB as vital differential microorganisms during Daqu fermentation. The correlation co-occurrence network constructed based on Spearman's rank correlation coefficients showed aggregation of LAB and Daqumicroorganisms, indicating LAB’s significant position in influencing the microbial community structure. LAB showed negative correlations with Bacillus, Saccharopolyspora, and Thermoactinomycesbut positive correlations with Issatchenkia, Candida, Acetobacter, and Gluconobacter. The predicted genes of LAB enriched 20 functional pathways during Daqu fermentation, including Biosynthesis of amino acids, Alanine, aspartate and glutamate metabolism, Valine, leucine and isoleucine biosynthesis and Starch and sucrose metabolism, which suggested that LAB had the functions of polysaccharide metabolism and amino acid biosynthesis.
CONCLUSION: Thus, the study indicates that LAB is important in determining the composition and function of Daqu microorganisms，and also suggests that LAB are closely related to the production of nitrogenous flavor substances in Daqu. The study provides a foundation for further exploring the function of LAB and the regulation of Daqu quality.
... Acetic acid is a weak organic acid harmful to most bacteria, even at concentrations as low as 0.5 wt%. Acetic acid, among other harmful effects, leads to a drop in intracellular pH and the disruption of some metabolic chains by acetic acid anion [37]. Figure 3B represents the bacteria cells after incubation in acetic acid. ...
Detecting bacteria—Listeria monocytogenes—is an essential healthcare and food industry issue. The objective of the current study was to apply platinum (Pt) and screen-printed carbon (SPCE) electrodes modified by molecularly imprinted polymer (MIP) in the design of an electrochemical sensor for the detection of Listeria monocytogenes. A sequence of potential pulses was used to perform the electrochemical deposition of the non-imprinted polypyrrole (NIP-Ppy) layer and Listeria monocytogenes-imprinted polypyrrole (MIP-Ppy) layer over SPCE and Pt electrodes. The bacteria were removed by incubating Ppy-modified electrodes in different extraction solutions (sulphuric acid, acetic acid, L-lysine, and trypsin) to determine the most efficient solution for extraction and to obtain a more sensitive and repeatable design of the sensor. The performance of MIP-Ppy- and NIP-Ppy-modified electrodes was evaluated by pulsed amperometric detection (PAD). According to the results of this research, it can be assumed that the most effective MIP-Ppy/SPCE sensor can be designed by removing bacteria with the proteolytic enzyme trypsin. The LOD and LOQ of the MIP-Ppy/SPCE were 70 CFU/mL and 210 CFU/mL, respectively, with a linear range from 300 to 6700 CFU/mL.
... Acid stress is often encountered by industrial microorganisms since organic acids are generated as either products or byproducts during multiple fermentation processes (Guan et al. 2017). Protonated acids may enter cells and then dissociate into protons and the corresponding ions, which leads to an increase in intracellular acidity and accelerates metabolic disorders in cells (Trček et al. 2015). In response, microorganisms have evolved a variety of strategies to resist acid stress for survival. ...
Streptomyces albulus is a well-established cell factory for ε-poly-L-lysine (ε-PL) production. It has been reported that ε-PL biosynthesis is strictly regulated by pH and that ε-PL can accumulate at approximately pH 4.0, which is outside of the general pH range for natural product production by Streptomyces species. However, how S. albulus responds to low pH is not clear. In this study, we attempted to explore the response of S. albulus to low-pH stress at the physiological and global gene transcription levels. At the physiological level, S. albulus maintained intracellular pH homeostasis at ~pH 7.5, increased the unsaturated fatty acid ratio, extended the fatty acid chain length, enhanced ATP accumulation, increased H⁺-ATPase activity, and accumulated the basic amino acids L-lysine and L-arginine. At the global gene transcription level, carbohydrate metabolism, oxidative phosphorylation, macromolecule protection and repair, and the acid tolerance system were found to be involved in combating low-pH stress. Finally, we preliminarily evaluated the effect of the acid tolerance system and cell membrane fatty acid synthesis on low-pH tolerance via gene manipulation. This work provides new insight into the adaptation mechanism of Streptomyces to low-pH stress and a new opportunity for constructing robust S. albulus strains for ε-PL production.
Key points
• S. albulus consistently remained pHiat ~7.4 regardless of the environmental pH.
• S. albulus combats low-pH stress by modulating lipid composition of cell membrane.
• Overexpression of cfa in S. albulus could improve low-pH tolerance and ɛ-PL titer.
Graphical abstract
... Because weak organic acids are lipophilic and can easily enter the plasma membrane, as well as intracellular acidification, they are more inhibitory than strong mineral acids. 31,32 ...
A novel organic acid-modified starch and carboxymethyl cellulose (CMC) based films plasticized with glycerol were prepared from unconventional tikhur starch (Curcuma angustifolia) by solution casting. Wet milling was used in the laboratory to extract starch from the tikhur rhizome. Carboxymethyl cellulose, at a concentration of (0.2 g−1 starch dry basis) was blended with the starch to improve its film-forming properties. Three different treatments with varying organic acids (lactic, citric, and acetic acid) with a concentration of 5% w/w of starch (2 ppm) in a film-forming solution were given. The effect of organic acid incorporation on the antibacterial, morphological, structural, thermal, and crystalline properties of developed films was studied. The minimum inhibitory concentration values of the three organic acids against gram-negative (E. coli) and gram-positive (S. aureus) bacteria were measured using the tube dilution method. The MIC results revealed that lactic acid and citric acid are effective against both gram-negative and gram-positive bacteria, while acetic acid showed more effectiveness against gram-negative bacteria (E. coli). MBC results revealed that organic acids have potent bactericidal activity. Citric acid resulted in higher inhibition for gram-positive bacteria (S. aureus) compared to gram-negative bacteria (E. coli.). While acetic acid showed higher inhibition for E. coli. than S. aureus. Lactic acid displayed similar inhibition against both S. aureus and E. coli. Among different organic acids, lactic acid incorporation resulted in a more homogeneous, transparent, and thermally stable film. As evidenced by the micrographs, the lactic acid incorporation resulted in a compact film structure without any visible cracks. While X-ray diffraction showed an increase in crystalline properties due to organic acid modification. In this study, it was indicated that modification with organic acids (polycarboxylic acids) effectively improved the overall properties of developed films depending on the type of organic acid used. The developed films have the potential to replace harmful synthetic films in food packaging.
Acinetobacter baumannii is a critical biofilm-forming pathogen that has presented great challenges in the clinic due to multidrug resistance. Thus, new methods of intervention are needed to control biofilm-associated infections. In this study, among three tested Lactobacillus species, Lactobacillus rhamnosus showed significant antimaturation and antiadherence effects against A. baumannii biofilm. Lactic acid (LA) and acetic acid (AA) were the most effective antibiofilm biosurfactants (BSs) produced by L. rhamnosus. This antibiofilm phenomenon produced by LA and AA was due to the strong bactericidal effect, which worked from very early time points, as determined by colony enumeration and confocal laser scanning microscope. The cell destruction of A. baumannii appeared in both the cell envelope and cytoplasm. A discontinuous cell envelope, the leakage of cell contents, and the increased extracellular activity of ATPase demonstrated the disruption of the cell membrane by LA and AA. These effects also demonstrated the occurrence of protein lysis. In addition, bacterial DNA interacted with and was damaged by LA and AA, resulting in significantly reduced expression of biofilm and DNA repair genes. The results highlight the possibility and importance of using probiotics in clinical prevention. Probiotics can be utilized as novel biocides to block and decrease biofilm formation and microbial contamination in medical equipment and during the treatment of infections. IMPORTANCE A. baumannii biofilm is a significant virulence factor that causes the biofilm colonization of invasive illnesses. Rising bacterial resistance to synthetic antimicrobials has prompted researchers to look at natural alternatives, such as probiotics and their derivatives. In this study, L. rhamnosus and its BSs (LA and AA) demonstrated remarkable antibiofilm and antimicrobial characteristics, with a significant inhibitory effect on A. baumannii. These effects were achieved by several mechanisms, including the disruption of the cell envelope membrane, protein lysis, reduced expression of biofilm-related genes, and destruction of bacterial DNA. The results provide support for the possibility of using probiotics and their derivatives in the clinical prevention and therapy of A. baumannii infections.
This study compared differences in physicochemical characteristics of the vinegar made by a mixed culture (MC) of Saccharomyces cerevisiae and Lactiplantibacillus plantarum and a pure culture (PC) of Saccharomyces cerevisiae. The fermentation process was monitored, and metabolomics analysis by Liquid Chromagraphy-Mass Spectrometry (LC-MS) was applied to the compositional differences between PC and MC vinegars, combined with quantification of organic acids, amino acids and B vitamins. A total of 71 differential metabolites including amino acids, organic acids and carbohydrates, and six possible key metabolic pathways were identified. MC enhanced the malic acid utilization and pyruvate acid metabolism during fermentation, increasing substrate-level phosphorylation, and supplying more energy for cellular metabolism. Higher acidity at the beginning of acetic acid fermentation, resulting from lactic acid production by Lactiplantibacillus plantarum in MC, suppressed the cellular metabolism and growth of Acetobacter pasteurianus, but enhanced its alcohol metabolism and acetic acid production in MC. MC vinegar contained more vitamin B, total flavonoids, total organic acids, amino acids and had a higher antioxidant capacity. MC enhanced the volatile substances, particularly ethyl lactate, ethyl caprate and ethyl caproate, which contributed to a stronger fruity aroma. These results indicated the mixed culture in alcoholic fermentation can effectively enhance the flavor and quality of apple cider vinegar.
Introduction:Transthoracic needle biopsy (TNB) is a common, safe and inexpensive procedure used in the diagnosis of intrathoracic lesions. Until today, there is still no study about the influence of standardized uptake value (SUVmax) in positron emission tomography/computed tomography (PET/CT) on the diagnostic success of TNB in intrathoracic lesions. We aimed to analyze the factors, one of which was the SUVmax value influencing the diagnosis success of CT-guided TNB in pulmonary, mediastinal and pleural lesions. Secondary aim was to investigate the predictive clinical factors of complications.Methods:A retrospective study of 403 patients who underwent CT-guided TNB at a reference chest diseases hospital between February 2019 and February 2021 was conducted. A pulmonologist had performed the procedure with a 20-gauge fine-needle (Spinal) or a 14-gauge automated needle (tru-cut). Data of pathology, microbiology and clinical follow-up of the patients were collected. A chi-square and Student’s t-test were used to evaluate the patient-related factors (gender and smoking), lesion-related factors (type, side, location, size, presence of necrosis observed by CT, SUVmax value in PET/CT), and procedure factor (type of needle) on the diagnostic success. Additionally, associations between clinical characteristics of patients and the complications were assessed.Results:A total of 403 patients underwent CT-guided TNB were enrolled and overall success was 70% (284/403). Smoking history (75% vs 43%, p=0.02) was predictor for diagnostic success. Lesion size and SUVmax value were significantly high in diagnosed patients (for both p
Environmental factors play an important role in contributing to intricate compositional dynamics and volatile metabolites in food fermentation. However, our understanding of which and how environmental factors affect volatile metabolites during sesame flavor-type baijiu fermentation is poor. Here, we examined the effects of environmental factors on the bacterial and fungal community to determine how changes in representative factors impact the microbial structure, diversity, and volatile metabolites in three fermentations. Results showed that bacterial community (ANOSIM: R = 0.79, P = 0.001), fungal community (ANOSIM: R = 0.65, P = 0.001), and volatile metabolites (ANOSIM: R = 0.84, P = 0.001) were significantly different in three fermentations. Acidity, ethanol, and moisture negatively impacted bacterial composition and diversity (P < 0.05). The fungal diversity and structure were positively and significantly affected by acidity (path coefficient, b = 0.54 for diversity, b = 0.35 for structure, P < 0.05). The fungal community rather than the bacterial community was the strongest driver of volatile metabolites. Fungal structure and diversity were equally important for the composition and content of volatile metabolites (structure: b = 0.50, diversity: b = 0.56, P < 0.05). 66 % of variations in volatile metabolites could be explained. Altogether these results indicated that acidity strongly drove volatile metabolites by modulating fungal structure and diversity. This work provides insights into managing volatile metabolites by regulating initial acidity in sesame flavor-type baijiu fermentation.
Strain engineering for industrial production requires the improvement of tolerance to multiple unfavorable conditions. Here, we report global regulator libraries based on the CRISPR‐enabled trackable genome engineering (CREATE) to engineer tolerance against multiple inhibitors in Escherichia coli. Deep mutagenesis libraries were rationally designed, and constructed to target 34,340 mutations across 23 global regulators. 69 specific mutations respectively conferred tolerance to acetate, NaCl, furfural, and high temperature were isolated, confirmed, and evaluated. Among them, 31 novel reconstructed mutations exhibited better tolerance to the corresponding inhibitors. The most dramatic mutation CRP‐E182D conferred high cross‐tolerance to acetate, NaCl, and isobutanol. According to the reconstruction and transcriptome analysis, the tolerance of mutant strains to acetate is a complex whole‐cell process. In particular, it was confirmed for the first time that upregulation of ilvA and nadA‐pnuC, deletion of potF or the sRNA sgrS, and adding exogenous valine or isoleucine significantly increased acetate tolerance. This article is protected by copyright. All rights reserved.
The presence of acetate exceeding 5 g/L is a major concern during E. coli fermentation due to its inhibitory effect on cell growth, thereby limiting high-density cell culture and recombinant protein production. Hence, engineered E. coli strains with enhanced acetate tolerance would be valuable for these bioprocesses. In this work, the acetate tolerance of E. coli was much improved by rewiring its global regulator cAMP receptor protein (CRP), which is reported to regulate 444 genes. Error-prone PCR method was employed to modify crp and the mutagenesis libraries (~3×10(6)) were subjected to M9 minimal medium supplemented with 5-10 g/L sodium acetate for selection. Mutant A2 (D138Y) was isolated and its growth rate in 15 g/L sodium acetate was found to be 0.083 h(-1), much higher than that of the control (0.016 h(-1)). Real-time PCR analysis via OpenArray(®) system revealed that over 400 CRP-regulated genes were differentially expressed in A2 with or without acetate stress, including those involved in the TCA cycle, phosphotransferase system, etc. Eight genes were chosen for overexpression and the overexpression of uxaB was found to lead to E. coli acetate sensitivity.
For the isolation of acetic acid bacteria from the »seed vinegar« culture, laboratory vinegar production was initiated with a culture for spirit vinegar production. The second and the third cycles of the process were followed. Samples for the isolation of acetic acid bacteria from the bioprocess liquid were taken four times during each cycle. A total of forty-seven strains were isolated. They were phenotyped as Acetobacter sp. Forty-one of them were identified as A. europaeus. Differentiation among 13 strains of A. europaeus and four other Acetobacter strains not identified as A. europaeus was done by random amplification of polymorphic DNA (RAPD) analysis. Seven different RAPD profiles were identified. From this it was assumed that in spirit vinegar genotypically different strains of acetic acid bacteria were present.
The pH and organic acid content of foods are two significant factors that may determine not only the types of organisms that will survive during storage but also their activities as spoilage organisms. In general, growth of important food-spoilage and food-poisoning organisms occurs over the range pH 4–8, whereas spoilage yeasts and molds are able to grow and survive at lower pH. Although organisms do exist that grow outside these pH ranges they have not in the past represented a significant problem in food spoilage. The pH of a food can be lowered artificially by the addition of significant amounts of acid: for example, acetic, citric, and lactic acids are often added to foods to lower the pH and in this way to limit microbial growth. This approach may be complemented by the addition of low concentrations of specific lipid-soluble weak acids, for example, benzoic and sorbic acids. The combined effect of a low pH plus a high weak-acid concentration leads to acidification of the cytoplasm, which is usually sufficient to restrict microbial growth, but may also have other specific effects on cell activity. Over recent years, greater insights have emerged into the mechanisms employed by yeasts and bacteria to counter acid stress.
Phospholipids of Gluconobacter cerinus were investigated. The cells of this bacterium were found to have cardiolipin, phosphatidylglycerol, phosphatidylethanolamine and phosphatidylcholine together with three ornithine-containing lipids. Phosphatidylcholine was the most abundant of these phospholipids, and calculated at 31.0% of total phospholipids in the cells at the late logarithmic phase. The fatty acid of this lipid was composed of palmitate, stearate and vaccenate, and there were no remarkable differences in quality among fatty acids of these four phospholipids. The phosphatidylcholine was also found in other acetic acid bacteria.