Bacteria Cells Research Articles

Molecular basis for curvature formation in SepF polymerization

The self-assembly of proteins into curved structures plays an important role in many cellular processes. One good example of this phenomenon is observed in the septum-forming protein (SepF), which forms polymerized structures with uniform curvatures. SepF is essential for regulating the thickness of the septum during bacteria cell division. In Bacillus subtilis, SepF polymerization involves two distinct interfaces, the β-β and α-α interfaces, which define the assembly unit and contact interfaces, respectively. However, the mechanism of curvature formation in this step is not yet fully understood. In this study, we employed solid-state NMR (SSNMR) to compare the structures of cyclic wild-type SepF assemblies with linear assemblies resulting from a mutation of G137 on the β-β interface. Our results demonstrate that while the sequence differences arise from the internal assembly unit, the dramatic changes in the shape of the assemblies depend on the α-α interface between the units. We further provide atomic-level insights into how the angular variation of the α2 helix on the α-α interface affects the curvature of the assemblies, using a combination of SSNMR, cryo-electron microscopy, and simulation methods. Our findings shed light on the shape control of protein assemblies and emphasize the importance of interhelical contacts in retaining curvature.

Development of chitosan/carrageenan macrobeads for encapsulation of Paenibacillus polymyxa and its biocontrol efficiency against clubroot disease in Brassica crops

Clubroot, caused by the obligate parasite Plasmodiophora brassicae, is one of the most important diseases of brassicas. The antagonistic bacterium Paenibacillus polymyxa ZF129 can suppress clubroot while its effectiveness is often unstable. To control clubroot more effectively, the macrobeads for controlled release of ZF129 were prepared using microencapsulation technology. Macrobeads with various ratios of chitosan (2 % w/w): carrageenan (0.3 % w/v) were prepared by an ionotropic gelation method and the bacteria ZF129 was loaded into macrobeads. The 1:1 chitosan: carrageenan showed the maximum swelling ratio (634 %), and the maximum survival rate (61.52 ± 1.12 %) after freeze-drying. Fourier transform infrared revealed the electrostatic interactions between chitosan and carrageenan. The macrobeads can efficiently release ZF129 strains into phosphate buffer solution and reach equilibrium in 48 h. The maximum number of bacteria cells to be released in the soil was observed after 25–30 days. The control efficacy of ZF129 macrobeads (chitosan: carrageenan, 1:1) and ZF129 culture against clubroot disease was 76.33 ± 3.65 % and 59.76 ± 4.43 % in greenhouse experiments, respectively and the control efficacy was calculated as 60.74 ± 5.00 % for ZF129 macrobeads and 40.94 ± 4.05 % for ZF129 culture under field experiments, respectively. The ZF129 macrobeads had significant growth-promoting effects on pak choi and Chinese cabbage. The encapsulation method described in this study is a prudent approach toward efficient biopesticides utilization with reduced environmental implications.

What happens to Bifidobacterium adolescentis and Bifidobacterium longum ssp. longum in an experimental environment with eukaryotic cells?

BackgroundThe impact of probiotic strains on host health is widely known. The available studies on the interaction between bacteria and the host are focused on the changes induced by bacteria in the host mainly. The studies determining the changes that occurred in the bacteria cells are in the minority. Within this paper, we determined what happens to the selected Bifidobacterium adolescentis and Bifidobacterium longum ssp. longum in an experimental environment with the intestinal epithelial layer. For this purpose, we tested the bacteria cells' viability, redox activity, membrane potential and enzymatic activity in different environments, including CaCo-2/HT-29 co-culture, cell culture medium, presence of inflammatory inductor (TNF-α) and oxygen.ResultsWe indicated that the external milieu impacts the viability and vitality of bacteria. Bifidobacterium adolescentis decrease the size of the live population in the cell culture medium with and without TNF-α (p < 0.001 and p < 0.01 respectively). In contrast, Bifidobacterium longum ssp. longum significantly increased survivability in contact with the eukaryotic cells and cell culture medium (p < 0.001). Bifidobacterium adolescentis showed significant changes in membrane potential, which was decreased in the presence of eukaryotic cells (p < 0.01), eukaryotic cells in an inflammatory state (p < 0.01), cell culture medium (p < 0.01) and cell culture medium with TNF-α (p < 0.05). In contrast, Bifidobacterium longum ssp. longum did not modulate membrane potential. Instead, bacteria significantly decreased the redox activity in response to milieus such as eukaryotic cells presence, inflamed eukaryotic cells as well as the culture medium (p < 0.001). The redox activity was significantly different in the cells culture medium vs the presence of eukaryotic cells (p < 0.001). The ability to β-galactosidase production was different for selected strains: Bifidobacterium longum ssp. longum indicated 91.5% of positive cells, whereas Bifidobacterium adolescentis 4.34% only. Both strains significantly reduced the enzyme production in contact with the eukaryotic milieu but not in the cell culture media.ConclusionThe environmental-induced changes may shape the probiotic properties of bacterial strains. It seems that the knowledge of the sensitivity of bacteria to the external environment may help to select the most promising probiotic strains, reduce research costs, and contribute to greater reproducibility of the obtained probiotic effects.

Bacteria-based Sodium Alginate Formulation to Control Toxic Microcystis Blooms

Cyanobacterial bloom formation in freshwaters is a major socio-economic and health concern across the globe. Presently used chemical and physical control strategies are inefficient in complete removal of blooms and chemical application often lead to secondary pollution in water. Hence, the current study aimed to develop a bacteria-based formulation to control toxic bloom-forming Microcystis aeruginosa in freshwaters. Two bacterial strains, Exiguobacterium acetylicum and Pseudomonas previously isolated from Sri Lankan freshwaters and characterized for M. aeruginosa cell lysis and their microcystin (MC) toxin degradation were used in the study. Initially, suitability of bacterial strains to develop into solid formulations was evaluated through biofilm formation, antibiotic sensitivity and strain compatibility. Both strains formed biofilms on the surface of microtiter plates indicating their ability to attach and colonize on solid surfaces. The multiple antibiotic resistance indices for both strains were below the threshold risk level (0.2) against the seven tested classes of antibiotics. This result indicates that there is a low risk in introducing these bacterial strains to the natural environment. Further, plate assay showed that the two strains were compatible to stay together showing no antagonistic effect on the growth of each other. Having fulfilled all three criteria tested, the two strains were immobilized into beads (~5 mm) prepared from sodium alginate at 1:1 ratio of 1×108 cells/mL bacterial inoculum. Different weights (1.0, 2.0, 3.0 g) of bacteria-immobilized beads were enclosed in sachets made with Cambrella synthetic fabric. They were introduced to M. aeruginosa grown in BG-11 medium (OD=0.2, 730 nm) and kept at 26 °C. During incubation, growth stimulation of M. aeruginosa was visually observed in 1.0 and 2.0 g beads-containing cultures, whereas, gradual discoloration of colonies was observed with 3.0 g of beads. Microscopic observations also proved complete disintegration of M. aeruginosa colonies and lysis of cells in discolored cultures. After 15 days, M. aeruginosa cell lysis was estimated as a measure of chlorophyll degradation. The highest (8.4%) cell lysis was observed in cultures containing 3.0 g of beads. This indicates that bacteria cell lysis activity depends on the load of bacteria-immobilized beads. Degradation of MC toxin was estimated by enzyme-linked immunosorbent assay. The highest (57.6%) MC degradation was observed in cultures with 3.0 g of beads after 15 days of incubation. The efficiency of bacteria release from beads was tested by placing sachets in sterilized water. At 15 days, bacterial count in water was x107 CFU/mL, with all loadings of immobilized-bacteria indicating highly efficient bacteria release from the formulation. In conclusion, this study highlights that bacteria-based sodium alginate formulations can be made as a source of inoculum to control M. aeruginosa growth and MC degradation. &#x0D; Keywords: Bacteria-based formulations, Cell lysis, Microcystis blooms, Microcystin

Whole-cell bacterial biosensor applied to identify the presence of Thaumatotibia leucotreta larva in citrus fruits by volatile sensing

The false codling moth (FCM), Thaumatotibia leucotreta (Meyrick), is a major pest of citrus fruits and other plants and is defined as a quarantine pest in several countries. Supervision and visual monitoring are crucial for the detection of eggs and larvae. There is a growing need to implement alternative pest control strategies and early detection methodologies in the packing house before shipping. Biosensors may provide a better-suited solution, as they are portable, do not require expensive equipment, and may be used with simple methodologies. In this study, a whole-cell bacterial biosensor was developed to identify the presence of Thaumatotibia leucotreta larvae in citrus fruits. The volatile organic compounds (VOCs) released by the larvae were identified (n = 11), which were not previously tested. The change in the VOCs composition in infected and uninfected mandarin (Citrus reticulata) was examined using gas chromatography-mass spectrometry (GC-MS) and the bioluminescent bacterial panel. During this work, two fixation methods (alginate beads and tablets) were tested for the bioluminescent bacteria cells, and tablets were preferred due to their higher uniformity and consistency of the light signal from the cells. Furthermore, the response of the bacteria to the changes in the concentration of VOCs as a result of a change in the amount of the larvae was examined (larvae n = 3, 6, or 9). In addition, in order to demonstrate the ability of the whole-cell biosensor to detect the presence of larva within the fruit, infected and uninfected mandarins were tested on the 14th day of larval hatching. The results showed that the bioluminescent bacteria could differentiate between mandarins with and without larva. The differences in the light signals (Induction Factor (IF)) between the mandarin fruits can be seen in the results with the larva (cytotoxicity IF = 3.046, genotoxicity IF = 2.643, and quorum sensing IF = 1.954) and without (cytotoxicity IF = 1.589, genotoxicity IF = 1.149 and quorum sensing IF = 1.205). The findings of this study make it possible to further develop a prototype using bioluminescent bacteria in order to detect pests of citrus fruits. The early detection of infected fruits will allow their filtering from the crop and therefore reduce the damage to additional fruits. Even one larva can invalidate the whole stock and the plot for future export.

Complementary supramolecular drug associates in perfecting the multidrug therapy against multidrug resistant bacteria.

The inappropriate and inconsistent use of antibiotics in combating multidrug-resistant bacteria exacerbates their drug resistance through a few distinct pathways. Firstly, these bacteria can accumulate multiple genes, each conferring resistance to a specific drug, within a single cell. This accumulation usually takes place on resistance plasmids (R). Secondly, multidrug resistance can arise from the heightened expression of genes encoding multidrug efflux pumps, which expel a broad spectrum of drugs from the bacterial cells. Additionally, bacteria can also eliminate or destroy antibiotic molecules by modifying enzymes or cell walls and removing porins. A significant limitation of traditional multidrug therapy lies in its inability to guarantee the simultaneous delivery of various drug molecules to a specific bacterial cell, thereby fostering incremental drug resistance in either of these paths. Consequently, this approach prolongs the treatment duration. Rather than using a biologically unimportant coformer in forming cocrystals, another drug molecule can be selected either for protecting another drug molecule or, can be selected for its complementary activities to kill a bacteria cell synergistically. The development of a multidrug cocrystal not only improves tabletability and plasticity but also enables the simultaneous delivery of multiple drugs to a specific bacterial cell, philosophically perfecting multidrug therapy. By adhering to the fundamental tenets of multidrug therapy, the synergistic effects of these drug molecules can effectively eradicate bacteria, even before they have the chance to develop resistance. This approach has the potential to shorten treatment periods, reduce costs, and mitigate drug resistance. Herein, four hypotheses are presented to create complementary drug cocrystals capable of simultaneously reaching bacterial cells, effectively destroying them before multidrug resistance can develop. The ongoing surge in the development of novel drugs provides another opportunity in the fight against bacteria that are constantly gaining resistance to existing treatments. This endeavour holds the potential to combat a wide array of multidrug-resistant bacteria.

Reversing the Natural Drug Resistance of Gram-Negative Bacteria to Fusidic Acid via Forming Drug-Phospholipid Complex.

Drug resistance substantially compromises antibiotic therapy and poses a serious threat to public health. Fusidic acid (FA) is commonly used to treat staphylococcal infections, such as pneumonia, osteomyelitis and skin infections. However, Gram-negative bacteria have natural resistance to FA, which is almost restrained in cell membranes due to the strong interactions between FA and phospholipids. Herein, we aim to utilize the strong FA-phospholipid interaction to pre-form a complex of FA with the exogenous phospholipid. The FA, in the form of an FA-phospholipid complex (FA-PC), no longer interacts with the endogenous membrane phospholipids and thus can be delivered into bacteria cells successfully. We found that the water solubility of FA (5 µg/mL) was improved to 133 µg/mL by forming the FA-PC (molar ratio 1:1). Furthermore, upon incubation for 6 h, the FA-PC (20 µg/mL) caused a 99.9% viability loss of E. coli and 99.1% loss of P. aeruginosa, while free FA did not work. The morphology of the elongated bacteria cells after treatment with the FA-PC was demonstrated by SEM. The successful intracellular delivery was shown by confocal laser scanning microscopy in the form of coumarin 6-PC (C6-PC), where C6 served as a fluorescent probe. Interestingly, the antibacterial effect of the FA-PC was significantly compromised by adding extra phospholipid in the medium, indicating that there may be a phospholipid-based transmembrane transport mechanism underlying the intracellular delivery of the FA-PC. This is the first report regarding FA-PC formation and its successful reversing of Gram-negative bacteria resistance to FA, and it provides a platform to reverse transmembrane delivery-related drug resistance. The ready availability of phospholipid and the simple preparation allow it to have great potential for clinical use.

Rapid and Ultrasensitive Colorimetric Biosensors for Onsite Detection of Escherichia coli O157:H7 in Fluids.

This study presents a breakthrough in the field of onsite bacterial detection, offering an innovative, rapid, and ultrasensitive colorimetric biosensor for the detection of Escherichia coli (E. coli) O157:H7, using chemically modified melamine foam (MF). Different from conventional platforms, such as 96-well plates and fiber-based membranes, the modified MF features a macroporous reticulated three-dimensional (3D) framework structure, allowing fast and free movement of large biomolecules and bacteria cells through the MF structure in every direction and ensuring good accessibility of entire active binding sites of the framework structure with the target bacteria, which significantly increased sensitive and volume-responsive detection of whole-cell bacteria. The biosensing platform requires less than 1.5 h to complete the quantitative detection with a sensitivity of 10 cfu/mL, discernible by the naked eye, and an enhanced sensitivity of 5 cfu/mL with the help of a smartphone. Following a short enrichment period of 1 h, the sensitivity was further amplified to 2 cfu/mL. The biosensor material is volume responsive, making the biosensing platform sensitivity increase as the volume of the sample increases, and is highly suitable for testing large-volume fluid samples. This novel material paves the way for the development of volume-flexible biosensing platforms for the record-fast, onsite, selective, and ultrasensitive detection of various pathogenic bacteria in real-world applications.

Manipulation of Surface Electrical Charge on Nanocomposite Membranes Confers Wide Spectrum Bactericidal Effects and Promotes Tissue Regeneration

AbstractUtilization of electro‐responsive biomaterials with antibacterial properties is advantageous for facilitating septic wound healing and tissue regeneration. However, the dose‐response effects of electrical stimuli from these materials against bacteria are not rigorously characterized, and achieving synergy of bactericidal and pro‐regenerative effects of biomaterials remains a major challenge. Here, a graded series of flexible BaTiO3/P(VDF‐TrFE) electroactive nanocomposite membranes (EMs) are developed with varying surface charge intensities, to serve as antibacterial dressing for septic wound healing. EMs display broad‐spectrum antibacterial effects against both Gram‐positive and Gram‐negative bacteria in a dose‐dependent manner, depending on the magnitude of their surface electrical potential. Mechanistically, the surface charge of EMs increase intracellular levels of reactive oxygen species within bacteria cells, which in turn caused oxidative damage to the bacterial membrane, thereby suppressing bacterial activity and biofilm formation. Moreover, in vivo studies demonstrated that EMs effectively inhibited S. aureus infection and accelerated wound healing in a mouse skin defect model, as well as ameliorated P. gingivalis‐mediated periodontal inflammation in a mouse periodontitis model. Hence, this study optimizes the antibacterial properties of electroactive materials and characterizes the dose‐response effects of surface electrical charge against bacteria, thus validating the therapeutic applications of electroactive biomaterials in combating bacterial infection.

Copper catalysis the synthesis of functionalized glycine-based sulphonamides: In silico and in vitro antibacterial studies

The synthesis of functionalized glycine-based sulphonamides via copper catalyzed N-arylation reaction and the in silico, in vitro antibacterial studies is reported. The procedure involved the initial synthesis of substituted p-toluenesulphonamides and substituted benzenesulphonamides by the reaction of glycine with p-toluenesulphonyl chloride and benzenesulphonyl chloride respectively in aqueous basic medium. The syn[1]thesized compounds were acetylated by reacting them with acetic anhydride and sodium acetate followed by acylation and amidation to yield amidated p-toluenesulphonamides and benzenesulphonamides respectively. Copper catalyzed N-arylation of the amidated products with aryltriolborates resulted in the synthesis of benzene, and 2-chlorophenyl derivatives of the amidated products. The synthesized compounds were characterized using FTIR, 1HNMR and elemental analysis and the spectra were in agreement with the assigned structures. The in silico antibacterial studies revealed that the compounds possess significant antibacterial potency in the respective bacteria cells and could be further employed as potential anti-bacterial agents. The in vitro antimicrobial study revealed that most of the synthesized compounds possess antibacterial activities.

고초균-젖산균의 순차적 복합 발효를 통한 복합 기능성 물질 함유 황칠나무 추출물의 생산

The production of poly-γ-glutamic acid (γ-PGA) and γ-aminobutyric acid (GABA) was optimized by serial fermentation of Dendropanax morbiferus extract (DME) using Bacillus subtilis HA and Lactobacillus plantarum KS2020. The 1st alkaline fermentation was performed on 60% DME including 2% glucose and 10% monosodium ʟ-glutamate (MSG) as a precursor. The 1st fermented DME had 57 mg% tyrosine. Consequently, the 2nd lactic acid fermentation for 5 days increased the tyrosine content of 106 mg%. The mucilage containing γ-PGA showed a high content of 3.50% on the first day of alkaline fermentation and then increased to 4.10% after 2 days. The precursor (MSG) remaining in the 1st fermented DME was efficiently converted to GABA by the 2nd lactic acid fermentation in the presence of 5% skim milk, 1.5% glucose and 0.5% yeast extract, resulting in the production of 18.29 mg/mL GABA. The viable cells of lactic acid bacteria increased and indicated 9.49 log CFU/mL on the fermentation for 5 days, and the acidity of co-fermented DME indicated the highest value of 1.55%. Conclusively, the serial fermented DME has multi-functional ingredients containing γ-PGA, GABA, peptides and probiotics.

Assessment of pulsed dielectrophoretic-field flow fractionation separation coupled with fibre-optic detection on a lab-on-chip as a technique to separate similar bacteria cells

Assessment of pulsed dielectrophoretic-field flow fractionation separation coupled with fibre-optic detection on a lab-on-chip as a technique to separate similar bacteria cells

DOP34 Deciphering immune-epithelial interactions in health and in Inflammatory Bowel Disease

Abstract Background Intestinal homeostasis is dependent on appropriate interactions between various compartments including immune, mesenchymal, neural, epithelial and bacteria cells. Disrupt of these interactions has been associated with the development of Inflammatory Bowel Disease (IBD). Using intestinal organoids as a starting point, we built complexity into this model by adding other cellular components to be able to study these interactions in health and disease. One of the key immune players in the intestine is Innate Lymphoid Cells (ILC). ILC precursors (ILCP) migrate to mucosa where they mature, promote homeostasis, and provide a potent, antigen-non-specific sources of cytokines. Deciphering what local stimuli drive the final stages of ILCP maturation in these tissues remains a pressing question, as ILC frequencies can become dysregulated in IBD. Methods Here, we develop and use co-cultures gut organoids with Innate Lymphoid Cells precursors (ILCP) and with mature intestinal Innate Lymphoid Cells (ILC). Results Harnessing these versatile models, we demonstrate that epithelial cells provide a complex niche capable of supporting the final maturation of all subsets including ILC1, ILC2, ILC3 and Natural Killer (NK) cells, Notably, organoid identity was sufficient to robustly recapitulate tissue-specific ILC imprints and frequencies, even in the absence of microbial stimuli, other cell types, or cytokine supplementation. In addition, we show that human gut ILC1 drive intestinal and extracellular matrix remodelling through production of TGFβ. This indicates the potential impact of ILC1 accumulation in IBD patients in driving intestinal cancer and fibrosis, two sequelae of IBD. Finally, we identify a new module of interaction between ILCs and the intestinal epithelium: goblet cells provide Notch ligands that are essential for maintenance of NC3R+ ILC3 and for their production of IL-22. In turn, ILC3 drive intestinal epithelial cells towards a secretory phenotype. Conclusion Taken together, our work provides unprecedented insight into in situ ILC maturation, which we show to be driven by epithelial signals, and into ILC function through deciphering new modules of ILC-epithelial interaction in health and IBD.

The species and physiological diversity of Bifidobacterium genus in Gallus gallus domesticus are influenced by feeding model and niche adaptations.

Species diversity of the Bifidobacterium genus was scarcely explored in different rearing systems of poultry. The aim of the study was to isolate intestinal species and compare their physiological and traits for adaptation to the avian intestinal niche. Fourteen strains isolated from chickens of intensive rearing farms and free-range hens, were identified by 16S rDNA sequencing, rep-PCR fingerprinting, and carbohydrates fermentation. Strains belonged to species Bifidobacterium pseudolongum subsp. pseudolongum and subsp. globosum, B. pullorum, B. animalis subsp lactis, B. boum, B. thermacidophilum subsp. thermacidophilum and B. thermophilum. One strain of B. animalis and B.pullorum, and two of B. pseudolongum subsp. pseudolongum were obtained from chicks, while the others were from free-range adult hens. Growth (in MRSc) at the poultry physiological temperature, acids production in caecal water with raffinose (rCW), ex vivo adhesion (%) to avian intestinal epithelial cells (IEC), and auto-aggregation (%) were used for discrimination inter- and intra-specific. Significantly different acetic and lactic acids production and growth temperatures were observed in strains of the same species/subspecies. Remarkable auto-aggregation capability was observed in B. thermacidophilum subsp. thermacidophilum LET 406 (40.2± 1.1%), while adhesion property was highlighted in B. pseudolongum subsp. pseudolongum LET 408 (65.30± 4.75% in jejunum; 46.05± 2.80 in ileum). Scanning Electronic Microscopy of the interaction IEC-LET 408 revealed an irregular bacterial surface exhibiting vesicle-like arrangements and filaments that formed a network among bacteria cells and with the epithelial cells, as possible adaptative response to promote its persistence in the gut. These finds will be valuable for bacterial supplements design intended to intensive rearing.

Assessment and Mitigation of Bacterial and Fungal Contamination in Refrigerator Waterlines

Three experiments were conducted on the recovery of microorganisms associated with refrigerator water lines. In the first experiment, between 2 and 3 logs CFU/ml were recovered from 100 different refrigerators’ residential water and ice lines. In a second experiment, plastic tubing segments cut from commercial refrigerator waterline systems were inoculated with the bacterium Pseudomonas aeruginosa and a fungal strain of Aspergillus flavus. Either 0.02% peracetic acid (PAA) exposure for 2 min or 0.03% PAA exposure for 1 min resulted in no recoverable bacteria cells, however, 0.25% PAA contact for 2 min was needed to eliminate detection of fungal cells. The third experiment tested sanitation of a water system inoculated with a mixed culture of P. aeruginosa and A flavus with a water filter cartridge filled with 3.5% PAA. The 3.5% concentration was determined using a computer simulation for mixing during a cleaning cycle that would yield a minimum concentration of at least 0.25% throughout the water system. After loading a water filter cartridge containing 3.5% PAA, allowing water to flow into the system for 15 s, and then allowing the system to sit for 5 min, no viable P. aeruginosa or A flavus cells were recovered up to 10 days after treatment.

Biological Function of Prophage-Related Gene Cluster ΔVpaChn25_RS25055~ΔVpaChn25_0714 of Vibrio parahaemolyticus CHN25.

Vibrio parahaemolyticus is the primary foodborne pathogen known to cause gastrointestinal infections in humans. Nevertheless, the molecular mechanisms of V. parahaemolyticus pathogenicity are not fully understood. Prophages carry virulence and antibiotic resistance genes commonly found in Vibrio populations, and they facilitate the spread of virulence and the emergence of pathogenic Vibrio strains. In this study, we characterized three such genes, VpaChn25_0713, VpaChn25_0714, and VpaChn25_RS25055, within the largest prophage gene cluster in V. parahaemolyticus CHN25. The deletion mutants ΔVpaChn25_RS25055, ΔVpaChn25_0713, ΔVpaChn25_0714, and ΔVpaChn25_RS25055-0713-0714 were derived with homologous recombination, and the complementary mutants ΔVpaChn25_0713-com, ΔVpaChn25_0714-com, ΔVpaChn25_RS25055-com, ΔVpaChn25_RS25055-0713-0714-com were also constructed. In the absence of the VpaChn25_RS25055, VpaChn25_0713, VpaChn25_0714, and VpaChn25_RS25055-0713-0714 genes, the mutants showed significant reductions in low-temperature survivability and biofilm formation (p < 0.001). The ΔVpaChn25_0713, ΔVpaChn25_RS25055, and ΔVpaChn25_RS25055-0713-0714 mutants were also significantly defective in swimming motility (p < 0.001). In the Caco-2 model, the above four mutants attenuated the cytotoxic effects of V. parahaemolyticus CHN25 on human intestinal epithelial cells (p < 0.01), especially the ΔVpaChn25_RS25055 and ΔVpaChn25_RS25055-0713-0714 mutants. Transcriptomic analysis showed that 15, 14, 8, and 11 metabolic pathways were changed in the ΔVpaChn25_RS25055, ΔVpaChn25_0713, ΔVpaChn25_0714, and ΔVpaChn25_RS25055-0713-0714 mutants, respectively. We labeled the VpaChn25_RS25055 gene with superfolder green fluorescent protein (sfGFP) and found it localized at both poles of the bacteria cell. In addition, we analyzed the evolutionary origins of the above genes. In summary, the prophage genes VpaChn25_0713, VpaChn25_0714, and VpaChn25_RS25055 enhance V. parahaemolyticus CHN25's survival in the environment and host. Our work improves the comprehension of the synergy between prophage-associated genes and the evolutionary process of V. parahaemolyticus.

Exploring the Synergy between Nano-Formulated Linezolid and Polymyxin B as a Gram-Negative Effective Antibiotic Delivery System Based on Mesoporous Silica Nanoparticles.

Antimicrobial resistance is a current silent pandemic that needs new types of antimicrobial agents different from the classic antibiotics that are known to lose efficiency over time. Encapsulation of antibiotics inside nano-delivery systems could be a promising, effective strategy that is able to delay the capability of pathogens to develop resistance mechanisms against antimicrobials. These systems can be adapted to deliver already discovered antibiotics to specific infection sites in a more successful way. Herein, mesoporous silica nanomaterials are used for an efficient delivery of a linezolid gram-positive antibiotic that acts synergistically with gram-negative antimicrobial polymyxin B. For this purpose, linezolid is encapsulated in the pores of the mesoporous silica, whose outer surface is coated with a polymyxin B membrane disruptor. The nanomaterial showed a good controlled-release performance in the presence of lipopolysaccharide, found in bacteria cell membranes, and the complete bacteria E. coli DH5α. The performed studies demonstrate that when the novel formulation is near bacteria, polymyxin B interacts with the cell membrane, thereby promoting its permeation. After this step, linezolid can easily penetrate the bacteria and act with efficacy to kill the microorganism. The nano-delivery system presents a highly increased antimicrobial efficacy against gram-negative bacteria, where the use of free linezolid is not effective, with a fractional inhibitory concentration index of 0.0063 for E. coli. Moreover, enhanced toxicity against gram-positive bacteria was confirmed thanks to the combination of both antibiotics in the same nanoparticles. Although this new nanomaterial should be further studied to reach clinical practice, the obtained results pave the way to the development of new nanoformulations which could help in the fight against bacterial infections.

Inactivation activity and mechanism of pulsed light against Alicyclobacillus acidoterrestris vegetative cells and spores in concentrated apple juice

Alicyclobacillus acidoterrestris has received much attention due to its unique thermo-acidophilic property and implication in the spoilage of pasteurized juices. The objective of this study was to evaluate the sterilization characteristics and mechanisms of pulsed light (PL) against A. acidoterrestris vegetative cells and spores in apple juice. The results indicated that bacteria cells in apple juice (8–20°Brix) can be completely inactivated within the fluence range of 20.25–47.25 J/cm2, which mainly depended on the soluble solids content (SSC) of juice, and the spores in apple juice (12°Brix) can be completely inactivated by PL with the fluence of 54.00 J/cm2. The PL treatment can significantly increase the leakage of reactive oxygen species (ROS) and proteins from cells and spores. Fluorescence studies of bacterial adenosine triphosphate (ATP) indicated that the loss of ATP was evident. Scanning electron microscopy and confocal laser scanning microscope presented that PL-treated cells or spores had serious morphological damage, which reduced the integrity of cell membrane and led to intracellular electrolyte leakage. In addition, there were no significant negative effects on total sugars, total acids, total phenols, pH value, SSC and soluble sugars, and organic acid content decreased slightly during the PL treatment. The contents of esters and acids in aroma components had a certain loss, while that of alcohols, aldehydes and ketones were increased. These results demonstrated that PL treatment can effectively inactivate the bacteria cells and spores in apple juice with little effect on its quality. This study provides an efficient method for the inactivation of A. acidoterrestris in fruit juice.

Design, biological characteristics, and antibacterial mechanism of high therapeutic index antimicrobial peptides with PRRP as central axis.

As the important components of biological innate immunity, antimicrobial peptides (AMPs) were found in a variety of organisms including insects, plants, animals, bacteria, fungi, etc. However, high hemolytic activity, high toxicity, and poor stability of natural AMPs hinder serious their application as therapeutic agents. To overcome these problems, in this study we use PRRP as a central axis, and peptides were designed based on the sequence template XRRXXRXPRRPXRXXRRX-NH2, where X represents a hydrophobic amino acid like Phe (F), Ile (I), Val (V), and Leu (L). The designed peptides LR18, FR18, and IR18 showed effective antimicrobial activity against some Gram-positive bacteria and Gram-negative bacteria, low cytotoxicity to mammalian cells, and had a tendency to form α-helical structures in membrane-mimetic environments. Among them, peptide LR18 (X: L) showed the highest geometric mean average treatment index (GMTI = 42.7) against Gram-negative bacteria, and FR18 (X: L) showed the highest GMTI (22.86) against Gram-positive bacteria. LR18 and FR18 also showed better salt, temperature, pH, and trypsin stability. LR18 and FR18 exert their antimicrobial effects mainly through destroying bacteria cell membrane. Briefly, peptide LR18 and FR18 have the potential to serve as a therapeutic agent to reduce antibiotic resistance owing to its high therapeutic index and great stability.

Uranium biomineralization by immobilized Chryseobacterium sp. strain PMSZPI cells for efficient uranium removal

Uranium (U) contamination is hazardous to human health and the environment owing to its radiotoxicity and chemical toxicity and needs immediate attention. In this study, the immobilized biomass of Chryseobacterium sp. strain PMSZPI isolated from U enriched site, was investigated for U(VI) biomineralization in batch and column set-up. Under batch mode, the fresh or lyophilized cells successfully entrapped in calcium alginate beads demonstrated effectual U precipitation under acid and alkaline conditions. The maximum removal was detected at pH 7 wherein ∼98–99% of uranium was precipitated from 1 mM uranyl carbonate solution loading ∼350 mg U/g of biomass within 24 h in the presence of organic phosphate substrate. The resulting uranyl phosphate precipitates within immobilized biomass loaded beads were observed by SEM-EDX and TEM while the formation of U biomineral was confirmed by FTIR and XRD. Retention of phosphatase activity without any loss of uranium precipitation ability was observed for alginate beads with lyophilized biomass stored for 90 d at 4 °C. Continuous flow through experiment with PMSZPI biomass immobilized in polyacrylamide gel exhibited U loading of 0.8 g U/g of biomass at pH 7 using 1 l of 1 mM uranyl solution. This investigation established the feasibility for the application of immobilized PMSZPI biomass for field studies. Environmental implicationUranium contamination is currently a serious environmental concern owing to anthropogenic activities and needs immediate attention. We have developed here a biotechnological method for successful uranium removal using immobilized cells of a uranium tolerant environmental bacterium, Chryseobacterium sp. strain PMSZPI isolated from U ore deposit via phosphatase enzyme mediated uranium precipitation. The ability of immobilized PMSZPI cells to precipitate U(VI) as long-term stable U phosphates under environmental conditions relevant for contaminated waters containing high concentrations of U that exerts toxicity for biological systems is explored here. The long term stability of the immobilized biomass without compromising its U removal capacity shows the relevance of the bioremediation strategy for uranium contamination proposed in this work.