Effect of Trehalose and Lactose Treatments on the Freeze-Drying Resistance of Lactic Acid Bacteria in High-Density Culture. - Related Documents




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601801.0000Effect of Trehalose and Lactose Treatments on the Freeze-Drying Resistance of Lactic Acid Bacteria in High-Density Culture. Freeze-drying is a commonly used method in commercial preparations of lactic acid bacteria. However, some bacteria are killed during the freeze-drying process. To overcome this, trehalose and lactose are often used as protective agents. Moreover, high-density culture is an efficient way to grow bacterial strains but creates a hypertonic growth environment. We evaluated the effects of trehalose and lactose, as a primary carbon source or as an additive in fermentation, on the freeze-drying survival of Lactobacillus fermentum FXJCJ6-1, Lactobacillus brevis 173-1-2, and Lactobacillus reuteri CCFM1040. Our results showed that L. fermentum FXJCJ6-1 accumulated but did not use intracellular trehalose in a hypertonic environment, which enhanced its freeze-drying resistance. Furthermore, genes that could transport trehalose were identified in this bacterium. In addition, both the lactose addition and lactose culture improved the freeze-drying survival of the bacterium. Further studies revealed that the added lactose might exert its protective effect by attaching to the cell surface, whereas lactose culture acted by reducing extracellular polysaccharide production and promoting the binding of the protectant to the cell membrane. The different mechanisms of lactose and trehalose in enhancing the freeze-drying resistance of bacteria identified in this study will help to elucidate the anti-freeze-drying mechanisms of other sugars in subsequent investigations.202236677339
15810.9996Homology- and cross-resistance of Lactobacillus plantarum to acid and osmotic stress and the influence of induction conditions on its proliferation by RNA-Seq. In this study, homology- and cross-resistance of Lactobacillus plantarum L1 and Lactobacillus plantarum L2 to acid and osmotic stress were investigated. Meanwhile, its proliferation mechanism was demonstrated by transcriptomic analysis using RNA sequencing. We found that the homologous-resistance and cross-resistance of L. plantarum L1 and L. plantarum L2 increased after acid and osmotic induction treatment by lactic acid and sodium lactate solution in advance, and the survival rate of live bacteria was improved. In addition, the count of viable bacteria of L. plantarum L2 significantly increased cultivated at a pH 5.0 with a 15% sodium lactate sublethal treatment, compared with the control group. Further study revealed that genes related to membrane transport, amino acid metabolism, nucleotide metabolism, and cell growth were significantly upregulated. These findings will contribute to promote high-density cell culture of starter cultures production in the fermented food industry.202133945164
471420.9995Screening and genome analysis of heat-resistant and antioxidant lactic acid bacteria from Holstein cow milk. BACKGROUND: Heat stress significantly impacts dairy cows, primarily through oxidative stress, which undermines their health. The problem is exacerbated by the ongoing global warming trend. Lactic acid bacteria (LAB) are safe, economical, and readily accessible options for enhancing the host's antioxidant defenses and preventing oxidative damage. They have been proven effective in alleviating heat stress-related damage, making them an excellent choice for protecting dairy cows from the adverse effects of heat stress. METHOD: In this study, five strains of LAB from Holstein cow milk (Lactobacillus plantarum L5, L14, L17, L19, L20) were evaluated for their heat resistance and antioxidant capacity by evaluating the growth characteristics and tolerance of the strains under high-temperature conditions, as well as their H(2)O(2) tolerance, free radical scavenging ability (DPPH, OH(-), ABTS), reducing ability, and EPS production ability. Furthermore, we employed Caco-2 cells to assess the adhesion rate of the strain, thereby confirming its ability to successfully colonize the host's intestinal tract and ensuring the effective execution of its probiotic functions. The strain with excellent heat resistance and antioxidant capacity was then subjected to genomic analysis to gain insight into the molecular mechanisms behind their heat resistance, antioxidant capacity, and safety. RESULTS: Among the two strains, Lactobacillus plantarum L19 emerges as a highly promising candidate. The strain exhibits robust growth even at high temperatures at 40°C and maintains a survival rate of 16.42% at the extreme temperature of 65°C. Furthermore, it demonstrates superior tolerance to hydrogen peroxide (27.3%), and possesses a notably higher free radical scavenging capacity with a high adhesion rate to Caco-2 cell (22.19%) compared to the other four strains tested. Genomic analysis revealed its' genome has 17 genes related to antioxidants and three genes related to heat resistance. Importantly, L19 lacks any resistance genes, ensuring its safety as a probiotic. CONCLUSION: The results imply that Lactobacillus plantarum L19 has the potential to serve as an effective food additive in mitigating damages associated with heat stress. This research offers a valuable reference for the prevention and management of heat stress in dairy cows, while also expanding the scope of applications for LAB derived from cow milk.202439611093
895430.9994Effect of biofilm formation by antimicrobial-resistant gram-negative bacteria in cold storage on survival in dairy processing lines. Antimicrobial-resistant gram-negative bacteria in dairy products can transfer antimicrobial resistance to gut microbiota in humans and can adversely impact the product quality. In this study, we aimed to investigate their distribution in dairy processing lines and evaluate biofilm formation and heat tolerance under dairy processing line-like conditions. Additionally, we compared the relative expression of general and heat stress-related genes as well as spoilage-related gene between biofilm and planktonic cells under consecutive stresses, similar to those in dairy processing lines. Most species of gram-negative bacteria isolated from five different dairy processing plants were resistant to one or more antimicrobials. Biofilm formation by the bacteria at 5 °C increased with the increase in exposure time. Moreover, cells in biofilms remained viable under heat treatment, whereas all planktonic cells of the selected strains died. The expression of heat-shock-related genes significantly increased with heat treatment in the biofilms but mostly decreased in the planktonic cells. Thus, biofilm formation under raw milk storage conditions may improve the tolerance of antimicrobial-resistant gram-negative bacteria to pasteurization, thereby increasing their persistence in dairy processing lines and products. Furthermore, the difference in response to heat stress between biofilm and planktonic cells may be attributed to the differential expression of heat stress-related genes. Therefore, this study contributes to the understanding of how gram-negative bacteria persist under consecutive stresses in dairy processing procedures and the potential mechanism underlying heat tolerance in biofilms.202336436412
15640.9994Bacterial Acid Resistance Toward Organic Weak Acid Revealed by RNA-Seq Transcriptomic Analysis in Acetobacter pasteurianus. Under extreme acidic environments, bacteria exploit several acid resistance (AR) mechanisms for enhancing their survival, which is concerned with several aspects, such as issues in human health and fermentation for acidic products. Currently, knowledge of bacterial AR mainly comes from the strong acid (such as hydrochloric acid) stresses, whereas AR mechanisms against organic weak acids (such as acetic acid), which are indeed encountered by bacteria, are less understood. Acetic acid bacteria (AAB), with the ability to produce acetic acid up to 20 g/100 mL, possess outstanding acetic acid tolerance, which is conferred by their unique AR mechanisms, including pyrroloquinoline quinine-dependent alcohol dehydrogenase, acetic acid assimilation and molecular chaperons. The distinguished AR of AAB toward acetic acid may provide a paradigm for research in bacterial AR against weak organic acids. In order to understand AAB's AR mechanism more holistically, omics approaches have been employed in the corresponding field. However, the currently reported transcriptomic study was processed under a low-acidity (1 g/100 mL) environment, which could not reflect the general conditions that AAB are usually faced with. This study performed RNA-Seq transcriptomic analysis investigating AR mechanisms in Acetobacter pasteurianus CGMCC 1.41, a widely used vinegar-brewing AAB strain, at different stages of fermentation, namely, under different acetic acid concentrations (from 0.6 to 6.03 g/100 mL). The results demonstrated the even and clustered genomic distribution of up- and down-regulated genes, respectively. Difference in AR between AAB and other microorganisms was supported by the down-regulation of urea degradation and trehalose synthesis-related genes in response to acetic acid. Detailed analysis reflected the role of ethanol respiration as the main energy source and the limited effect of acetic acid assimilation on AR during fermentation as well as the competition between ethanol respiratory chain and NADH, succinate dehydrogenase-based common respiratory chain. Molecular chaperons contribute to AR, too, but their regulatory mechanisms require further investigation. Moreover, pathways of glucose catabolism and fatty acid biosynthesis are also related to AR. Finally, 2-methylcitrate cycle was proposed as an AR mechanism in AAB for the first time. This study provides new insight into AR mechanisms of AAB, and it also indicates the existence of numerous undiscovered AR mechanisms.201931447789
675550.9993Impact of lead (Pb(2+)) on the growth and biological activity of Serratia marcescens selected for wastewater treatment and identification of its zntR gene-a metal efflux regulator. Microorganisms isolated from contaminated areas play an important role in bioremediation processes. They promote heavy metal removal from the environment by adsorbing ions onto the cell wall surface, accumulating them inside the cells, or reducing, complexing, or precipitating these substances in the environment. Microorganism-based bioremediation processes can be highly efficient, low-cost and have low environmental impact. Thus, the present study aimed to select Pb(2+)-resistant bacteria and evaluate the growth rate, biological activity, and the presence of genes associated with metal resistance. Serratia marcescens CCMA 1010, that was previously isolated from coffee processing wastewater, was selected since was able to growth in Pb(2+) concentrations of up to 4.0 mM. The growth rate and generation time did not differ from those of the control (without Pb(2+)), although biological activity decreased in the first hour of exposure to these ions and stabilized after this period. The presence of the zntR, zntA and pbrA genes was analysed, and only zntR was detected. The zntR gene encodes a protein responsible for regulating the production of ZntA, a transmembrane protein that facilitates Pb(2+) extrusion out of the cell. S. marcescens CCMA 1010 demonstrated a potential for use as bioindicator that has potential to be used in bioremediation processes due to its resistance to high concentrations of Pb(2+), ability to grow until 24 h of exposure, and possession of a gene that indicates the existence of mechanisms associated with resistance to lead (Pb(2+)).202336752862
15460.9993Classification of acetic acid bacteria and their acid resistant mechanism. Acetic acid bacteria (AAB) are obligate aerobic Gram-negative bacteria that are commonly used in vinegar fermentation because of their strong capacity for ethanol oxidation and acetic acid synthesis as well as their acid resistance. However, low biomass and low production rate due to acid stress are still major challenges that must be overcome in industrial processes. Although acid resistance in AAB is important to the production of high acidity vinegar, the acid resistance mechanisms of AAB have yet to be fully elucidated. In this study, we discuss the classification of AAB species and their metabolic processes and review potential acid resistance factors and acid resistance mechanisms in various strains. In addition, we analyze the quorum sensing systems of Komagataeibacter and Gluconacetobacter to provide new ideas for investigation of acid resistance mechanisms in AAB in the form of signaling pathways. The results presented herein will serve as an important reference for selective breeding of high acid resistance AAB and optimization of acetic acid fermentation processes.202133595734
15970.9993Putrescine production via the ornithine decarboxylation pathway improves the acid stress survival of Lactobacillus brevis and is part of a horizontally transferred acid resistance locus. Decarboxylation pathways are widespread among lactic acid bacteria; their physiological role is related to acid resistance through the regulation of the intracellular pH and to the production of metabolic energy via the generation of a proton motive force and its conversion into ATP. These pathways include, among others, biogenic amine (BA) production pathways. BA accumulation in foodstuffs is a health risk; thus, the study of the factors involved in their production is of major concern. The analysis of several lactic acid bacterial strains isolated from different environments, including fermented foods and beverages, revealed that the genes encoding these pathways are clustered on the chromosome, which suggests that these genes are part of a genetic hotspot related to acid stress resistance. Further attention was devoted to the ornithine decarboxylase pathway, which affords putrescine from ornithine. Studies were performed on three lactic acid bacteria belonging to different species. The ODC pathway was always shown to be involved in cytosolic pH alkalinisation and acid shock survival, which were observed to occur with a concomitant increase in putrescine production.201424495587
15580.9993RNA-Seq transcriptomic analysis reveals gene expression profiles of acetic acid bacteria under high-acidity submerged industrial fermentation process. Acetic acid bacteria (AAB) are Gram-negative obligate aerobics in Acetobacteraceae family. Producing acetic acid and brewing vinegars are one of the most important industrial applications of AAB, attributed to their outstanding ability to tolerate the corresponding stresses. Several unique acid resistance (AR) mechanisms in AAB have been revealed previously. However, their overall AR strategies are still less-comprehensively clarified. Consequently, omics analysis was widely performed for a better understanding of this field. Among them, transcriptome has recently obtained more and more attention. However, most currently reported transcriptomic studies were conducted under lab conditions and even in low-acidity environment, which may be unable to completely reflect the conditions that AAB confront under industrialized vinegar-brewing processes. In this study, we performed an RNA-Seq transcriptomic analysis concerning AAB's AR mechanisms during a continuous and periodical industrial submerged vinegar fermentation process, where a single AAB strain performed the fermentation and the acetic acid concentration fluctuated between ~8% and ~12%, the highest acidity as far we know for transcriptomic studies. Samples were directly taken from the initial (CK), mid, and final stages of the same period of the on-going fermentation. 16S rRNA sequence analysis indicated the participation of Komagataeibacter europaeus in the fermentation. Transcriptomic results demonstrated that more genes were downregulated than upregulated at both mid and final stages. Kyoto Encyclopedia of Genes and Genomes (KEGG) enrich analysis reflected that the upregulated genes mainly carried out tricarboxylic acid cycle and oxidative phosphorylation processes, probably implying a considerable role of acetic acid overoxidation in AR during fermentation. Besides, upregulation of riboflavin biosynthesis pathway and two NAD(+)-dependent succinate-semialdehyde dehydrogenase-coding genes suggested a critical role of succinate oxidation in AR. Meanwhile, downregulated genes were mainly ribosomal protein-coding ones, reflecting that the adverse impact on ribosomes initiates at the transcription level. However, it is ambiguous whether the downregulation is good for stress responding or it actually reflects the stress. Furthermore, we also assumed that the fermentation stages may have a greater effect on gene expression than acidity. Additionally, it is possible that some physiological alterations would affect the AR to a larger extent than changes in gene expression, which suggests the combination of molecular biology and physiology research will provide deeper insight into the AR mechanisms in AAB.202236246236
895190.9993Response mechanisms of resistance in L-form bacteria to different target antibiotics: Implications from oxidative stress to metabolism. Due to the specific action on bacterial cell wall, β-lactam antibiotics have gained widespread usage as they exhibit a high degree of specificity in targeting bacteria, but causing minimal toxicity to host cells. Under antibiotic pressure, bacteria may opt to shed their cell walls and transform into L-form state as a means to evade the antibiotic effects. In this study, we explored and identified diverse optimal conditions for both Gram-negative bacteria (E. coli DH5α (CTX)) and Gram-positive bacteria (B. subtilis ATCC6633), which were induced to L-form bacteria using lysozyme (0.5 ppm) and meropenem (64 ppm). Notably, when bacteria transformed into L-form state, both bacterial strains showed varying degrees of increased resistance to antibiotics polymyxin E, meropenem, rifampicin, and tetracycline. E. coli DH5α (CTX) exhibited the most significant enhancement in resistance to tetracycline, with a 128-fold increase, while B. subtilis ATCC6633 showed a 32-fold increase in resistance to tetracycline and polymyxin E. Furthermore, L-form bacteria maintained their normal metabolic activity, combined with enhanced oxidative stress, served as an adaptive strategy promoting the sustained survival of L-form bacteria. This study provided a theoretical basis for comprehending antibiotic resistance mechanisms, developing innovative treatment strategies, and confronting global antibiotic resistance challenges.202438735077
8679100.9993Metal accumulation in cell wall: a possible mechanism of cadmium resistance by Pseudomonas stutzeri. A heavy metal resistant strain, Pseudomonas stutzeri (MTCC 101) has been investigated for its cadmium tolerance properties along with its antibiotic resistance. The organism could tolerate cadmium up to 1,200 μg/mL with LD50 value 700 μg/mL. The gene(s) involved in such high resistance appear(s) to be induced in the presence of the metal. Increasing concentrations of cadmium successively prolonged the lag phase of growth with delayed attainment of the stationary phase. Transmission electron microscope and scanning electron microscope-energy dispersive analysis of X-ray spectroscope analysis showed cadmium adsorption on the bacterial surface with morphological distortion. Atomic absorption spectrometric study corroborated this data, showing highest cadmium accumulation in the cell wall fraction of the bacteria. Additionally, the cell wall fraction showed synthesis of new proteins when grown under metal stress.201323275974
8953110.9992Evolution of antibiotic resistance impacts optimal temperature and growth rate in Escherichia coli and Staphylococcus epidermidis. AIMS: Bacterial response to temperature changes can influence their pathogenicity to plants and humans. Changes in temperature can affect cellular and physiological responses in bacteria that can in turn affect the evolution and prevalence of antibiotic-resistance genes. Yet, how antibiotic-resistance genes influence microbial temperature response is poorly understood. METHODS AND RESULTS: We examined growth rates and physiological responses to temperature in two species-E. coli and Staph. epidermidis-after evolved resistance to 13 antibiotics. We found that evolved resistance results in species-, strain- and antibiotic-specific shifts in optimal temperature. When E. coli evolves resistance to nucleic acid and cell wall inhibitors, their optimal growth temperature decreases, and when Staph. epidermidis and E. coli evolve resistance to protein synthesis and their optimal temperature increases. Intriguingly, when Staph. epidermidis evolves resistance to Teicoplanin, fitness also increases in drug-free environments, independent of temperature response. CONCLUSION: Our results highlight how the complexity of antibiotic resistance is amplified when considering physiological responses to temperature. SIGNIFICANCE: Bacteria continuously respond to changing temperatures-whether through increased body temperature during fever, climate change or other factors. It is crucial to understand the interactions between antibiotic resistance and temperature.202236070219
157120.9992Analysis of proteins responsive to acetic acid in Acetobacter: molecular mechanisms conferring acetic acid resistance in acetic acid bacteria. Acetic acid bacteria are used for industrial vinegar production because of their remarkable ability to oxidize ethanol and high resistance to acetic acid. Although several molecular machineries responsible for acetic acid resistance in acetic acid bacteria have been reported, the entire mechanism that confers acetic acid resistance has not been completely understood. One of the promising methods to elucidate the entire mechanism is global analysis of proteins responsive to acetic acid by two-dimensional gel electrophoresis. Recently, two proteins whose production was greatly enhanced by acetic acid in Acetobacter aceti were identified to be aconitase and a putative ABC-transporter, respectively; furthermore, overexpression or disruption of the genes encoding these proteins affected acetic acid resistance in A. aceti, indicating that these proteins are involved in acetic acid resistance. Overexpression of each gene increased acetic acid resistance in Acetobacter, which resulted in an improvement in the productivity of acetic acid fermentation. Taken together, the results of the proteomic analysis and those of previous studies indicate that acetic acid resistance in acetic acid bacteria is conferred by several mechanisms. These findings also provide a clue to breed a strain having high resistance to acetic acid for vinegar fermentation.200817920150
8682130.9992Role of manganese superoxide dismutase (Mn-SOD) against Cr(III)-induced toxicity in bacteria. The toxicity of Cr(VI) was widely investigated, but the defense mechanism against Cr(III) in bacteria are seldom reported. Here, we found that Cr(III) inhibited bacterial growth and induced reactive oxygen species (ROS). After exposure to Cr(III), loss of sodA not only led to the excessive generation of ROS, but also enhanced the level of lipid peroxidation and reduced the GSH level, indicating that the deficiency of Mn-SOD decreased the bacterial resistance ability against Cr(III). The adverse effects of oxidative stress caused by Cr(III) could be recovered by the rescue of Mn-SOD in the sodA-deficient strain. Besides the oxidative stress, Cr(III) could cause the bacterial morphology variation, which was distinct between the wild-type and the sodA-deficient strains due to the differential expressions of Z-ring division genes. Moreover, Mn-SOD might prevent Cr(III) from oxidation on the bacterial surface by combining with Cr(III). Taken together, our results indicated that the Mn-SOD played a vital role in regulating the stress resistance, expression of cell division-related genes, bacterial morphology, and chemistry valence state of Cr. Our findings firstly provided a more in-depth understanding of Cr(III) toxicity and bacterial defense mechanism against Cr(III).202132781281
8949140.9992Potential Risk of Spreading Resistance Genes within Extracellular-DNA-Dependent Biofilms of Streptococcus mutans in Response to Cell Envelope Stress Induced by Sub-MICs of Bacitracin. Antibiotics are used to treat or prevent some types of bacterial infection. The inappropriate use of antibiotics unnecessarily promotes antibiotic resistance and increases resistant bacteria, and controlling these bacteria is difficult. While the emergence of drug-resistant bacteria is a serious problem, the behavior of drug-resistant bacteria is not fully understood. In this study, we investigated the behavior of Streptococcus mutans, a major etiological agent of dental caries that is resistant to bacitracin, which is a cell wall-targeting antibiotic, and focused on biofilm formation in the presence of bacitracin. S. mutans UA159 most strongly induced extracellular DNA (eDNA)-dependent biofilm formation in the presence of bacitracin at 1/8× MIC. The ΔmbrC and ΔmbrD mutant strains, which lack bacitracin resistance, also formed biofilms in the presence of bacitracin at 1/2× MIC. This difference between the wild type and the mutants was caused by the induction of atlA expression in the mid-log phase. We also revealed that certain rgp genes involved in the synthesis of rhamnose-glucose polysaccharide related to cell wall synthesis were downregulated by bacitracin. In addition, glucosyltransferase-I was also involved in eDNA-dependent biofilm formation. The biofilm led to increased transformation efficiencies and promoted horizontal gene transfer. Biofilms were also induced by ampicillin and vancomycin, antibiotics targeting cell wall synthesis, suggesting that cell envelope stress triggers biofilm formation. Therefore, the expression of the atlA and rgp genes is regulated by S. mutans, which forms eDNA-dependent biofilms, promoting horizontal gene transfer in response to cell envelope stress induced by sub-MICs of antibiotics.IMPORTANCE Antibiotics have been reported to induce biofilm formation in many bacteria at subinhibitory concentrations. Accordingly, it is conceivable that the MIC against drug-sensitive bacteria may promote biofilm formation of resistant bacteria. Since drug-resistant bacteria have spread, it is important to understand the behavior of resistant bacteria. Streptococcus mutans is bacitracin resistant, and the 1/8× MIC of bacitracin, which is a cell wall-targeted antibiotic, induced eDNA-dependent biofilm formation. The ΔmbrC and ΔmbrD strains, which are not resistant to bacitracin, also formed biofilms in the presence of bacitracin at 1/2× MIC, and biofilms of both the wild type and mutants promoted horizontal gene transfer. Another cell wall-targeted antibiotic, vancomycin, showed effects on biofilms and gene transfer similar to those of bacitracin. Thus, treatment with cell wall-targeted antibiotics may promote the spread of drug-resistant genes in biofilms. Therefore, the behavior of resistant bacteria in the presence of antibiotics at sub-MICs should be investigated when using antibiotics.202032532873
8680150.9992Environmental pH affects transcriptional responses to cadmium toxicity in Escherichia coli K-12 (MG1655). It has been widely reported that pH mediates cadmium toxicity to bacteria. We used a tripartite approach to investigate mechanisms by which pH affects cadmium toxicity that included analyses of: (1) growth kinetics, (2) global gene expression, and (3) cadmium speciation. Cadmium extended the lag phase at pH 7, but not at pH 5. DNA microarray analysis revealed that stress response genes including hdeA, otsA, and yjbJ were more highly expressed at pH 5 than at pH 7 after only 5 min of exposure to cadmium, suggesting that acidic pH more rapidly induced genes that confer cadmium resistance. In addition, genes involved in transport and many hypothetical genes were more highly expressed at pH 5 than at pH 7 in the presence of cadmium. Concentrations of two cadmium species, including one previously implicated in the mechanism by which pH mediates cadmium toxicity (CdOH+), increased with pH. Our data demonstrate that transcriptional responses of Escherichia coli to cadmium are substantially affected by pH and suggest that several stress response, transport, and hypothetical genes play roles in the mechanism by which pH mediates cadmium toxicity.200919220470
8303160.9992Spaceflight Modifies Escherichia coli Gene Expression in Response to Antibiotic Exposure and Reveals Role of Oxidative Stress Response. Bacteria grown in space experiments under microgravity conditions have been found to undergo unique physiological responses, ranging from modified cell morphology and growth dynamics to a putative increased tolerance to antibiotics. A common theory for this behavior is the loss of gravity-driven convection processes in the orbital environment, resulting in both reduction of extracellular nutrient availability and the accumulation of bacterial byproducts near the cell. To further characterize the responses, this study investigated the transcriptomic response of Escherichia coli to both microgravity and antibiotic concentration. E. coli was grown aboard International Space Station in the presence of increasing concentrations of the antibiotic gentamicin with identical ground controls conducted on Earth. Here we show that within 49 h of being cultured, E. coli adapted to grow at higher antibiotic concentrations in space compared to Earth, and demonstrated consistent changes in expression of 63 genes in response to an increase in drug concentration in both environments, including specific responses related to oxidative stress and starvation response. Additionally, we find 50 stress-response genes upregulated in response to the microgravity when compared directly to the equivalent concentration in the ground control. We conclude that the increased antibiotic tolerance in microgravity may be attributed not only to diminished transport processes, but also to a resultant antibiotic cross-resistance response conferred by an overlapping effect of stress response genes. Our data suggest that direct stresses of nutrient starvation and acid-shock conveyed by the microgravity environment can incidentally upregulate stress response pathways related to antibiotic stress and in doing so contribute to the increased antibiotic stress tolerance observed for bacteria in space experiments. These results provide insights into the ability of bacteria to adapt under extreme stress conditions and potential strategies to prevent antimicrobial-resistance in space and on Earth.201829615983
8216170.9992The Effect of glycocholic acid on the growth, membrane permeability, conjugation and antibiotic susceptibility of Enterobacteriaceae. INTRODUCTION: Glycocholic acid (GCA) is a steroid acid and one of the main glycine-conjugated bile components in mammalian bile, which is involved in the emulsification and absorption of fats and sterols. It is long-known that the amphipathic nature of bile acids enables them to interact with the lipid membrane of Gram-positive bacteria and act as potent antimicrobial compounds. Nevertheless, Gram-negative Enterobacteriaceae species inhabiting the intestinal tract of mammals are considered to be more bile-resistant compared to Gram-positive bacteria and are thought to tolerate high bile concentrations. RESULTS: Here, we show that 1-2% of GCA inhibit the growth of Enterobacteriaceae species, including E. coli, Salmonella enterica. Klebsiella spp., Citrobacter spp., and Raoultella spp. during their late logarithmic phase in liquid culture, but not in solid media. Despite their lipopolysaccharide membrane layer, we demonstrate that, in liquid, GCA increases permeability, changes the surface of the Enterobacteriaceae membrane, and compromises its integrity. These changes result in leakage of cytoplasmic proteins and enhancement of their susceptibility to antibiotics. Moreover, GCA significantly reduces bacterial motility, the frequency of bacterial conjugation and horizontal acquisition of antibiotic resistance genes. These phenotypes are associated with repression of flagellin (fliC) transcription and a sharp decrease in the occurrence of conjugative pili in the presence of glycocholic acid, respectively. DISCUSSION: Overall, these findings broaden the current understanding about bile resistance of Gram-negative bacteria and suggest that GCA can be used to inhibit bacterial growth, augment the activity of antimicrobial compounds and diminish acquisition and dissemination of antibiotic resistance genes by conjugation.202540256452
6305180.9992Antimicrobial genes from Allium sativum and Pinellia ternata revealed by a Bacillus subtilis expression system. Antimicrobial genes are found in all classes of life. To efficiently isolate these genes, we used Bacillus subtilis and Escherichia coli as target indicator bacteria and transformed them with cDNA libraries. Among thousands of expressed proteins, candidate proteins played antimicrobial roles from the inside of the indicator bacteria (internal effect), contributing to the sensitivity (much more sensitivity than the external effect from antimicrobial proteins working from outside of the cells) and the high throughput ability of screening. We found that B. subtilis is more efficient and reliable than E. coli. Using the B. subtilis expression system, we identified 19 novel, broad-spectrum antimicrobial genes. Proteins expressed by these genes were extracted and tested, exhibiting strong external antibacterial, antifungal and nematicidal activities. Furthermore, these newly isolated proteins could control plant diseases. Application of these proteins secreted by engineered B. subtilis in soil could inhibit the growth of pathogenic bacteria. These proteins are thermally stable and suitable for clinical medicine, as they exhibited no haemolytic activity. Based on our findings, we speculated that plant, animal and human pathogenic bacteria, fungi or even cancer cells might be taken as the indicator target cells for screening specific resistance genes.201830266995
6291190.9992Adaptive Resistance of Staphylococcus aureus to Cefquinome Sulfate in an In Vitro Pharmacokinetic Model with Transcriptomic Insights. Cefquinome sulfate has a strong killing effect against Staphylococcus aureus (S. aureus), but bacterial resistance has become increasingly widespread. Experiments were conducted to investigate the pattern of adaptive resistance of S. aureus to cefquinome sulfate under different dosage regimens by using pharmacokinetic-pharmacodynamics (PK-PD) modeling, and the adaptive-resistant bacteria in different states were screened and subjected to transcriptomic sequencing. The results showed that the minimum inhibitory concentration of Staphylococcus aureus under the action of cefquinome sulfate was 0.5 μg/mL, the anti-mutation concentration was 1.6 μg/mL, and the mutation selection window range was 0.5~1.6 μg/mL. In the in vitro pharmacokinetic model to simulate different dosing regimens in the animal body, there are certain rules for the emergence of adaptive drug-resistant bacteria: the intensity of bacterial resistance gradually increased with culture time, and the order of emergence was tolerant bacteria (TO) followed by persistent bacteria (PE) and finally resistant bacteria (RE). The sequence reflected the evolution of adaptive drug resistance. Transcriptome Gene Ontology (GO) analysis revealed that differentially expressed genes were involved in cellular respiration, energy derivation by oxidation of organic compounds, and oxidation-reduction processes. The differentially expressed genes identified functioned in the synthesis of cell membranes, cytoplasm, and intracellular parts. A Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis found that 65 genes were differentially expressed after cefquinome sulfate treatment, of which 35 genes were significantly upregulated and 30 genes were significantly downregulated. Five genes, sdhB, sdhA, pdhA, lpdA, and sucC, may be involved in network regulation. This study revealed the cross-regulation of multiple metabolic pathway networks and the targets of network regulation of S. aureus to produce adaptive drug resistance. The results will provide guidance for clinical drug use in animals infected with S. aureus.202540005696