D-Amino Acids Indirectly Inhibit Biofilm Formation in Bacillus subtilis by Interfering with Protein Synthesis

The soil bacterium Bacillus subtilis forms biofilms on surfaces and at air-liquid interfaces. It was previously reported that these biofilms disassemble late in their life cycle and that conditioned medium from late-stage biofilms inhibits biofilm formation. Such medium contained a mixture of d-leucine, d-methionine, d-tryptophan, and d-tyrosine and was reported to inhibit biofilm formation via the incorporation of these d-amino acids into the cell wall. Here, we show that l-amino acids were able to specifically reverse the inhibitory effects of their cognate d-amino acids. We also show that d-amino acids inhibited growth and the expression of biofilm matrix genes at concentrations that inhibit biofilm formation. Finally, we report that the strain routinely used to study biofilm formation has a mutation in the gene (dtd) encoding d-tyrosyl-tRNA deacylase, an enzyme that prevents the misincorporation of d-amino acids into protein in B. subtilis. When we repaired the dtd gene, B. subtilis became resistant to the biofilm-inhibitory effects of d-amino acids without losing the ability to incorporate at least one noncanonical d-amino acid, d-tryptophan, into the peptidoglycan peptide side chain. We conclude that the susceptibility of B. subtilis to the biofilm-inhibitory effects of d-amino acids is largely, if not entirely, due to their toxic effects on protein synthesis.Chemistry and Chemical Biolog

Vibrio gazogenes-dependent disruption of aflatoxin biosynthesis in Aspergillus flavus: the connection with endosomal uptake and hyphal morphogenesis

Aflatoxins, a family of fungal secondary metabolites, are toxic and carcinogenic compounds that pose an enormous threat to global food safety and agricultural sustainability. Specifically agricultural products in African, Southeast Asian and hot and humid regions of American countries suffer most damage from aflatoxin producing molds due to the ideal climate conditions promoting their growth. Our recent studies suggest that Vibrio gazogenes (Vg), an estuarine bacterium non-pathogenic to plants and humans, can significantly inhibit aflatoxin biosynthesis in the producers. In this study, we investigated the mechanism underlying Vg-dependent aflatoxin inhibition using the prominent aflatoxin producer, Aspergillus flavus. We show that aflatoxin inhibition upon Vg treatment was associated with fungal uptake of Vg-prodigiosin, a red pigment, which was consistently visible inside fungal hyphae during treatment. The association of prodigiosin with aflatoxin inhibition was further evident as Serratia marcescens, another prodigiosin producer, significantly inhibited aflatoxin, while non-producers like Escherichia coli, Staphylococcus aureus, Vibrio harveyi, and Vibrio fischeri did not. Also, pure prodigiosin significantly inhibited aflatoxin biosynthesis. Endocytosis inhibitors, filipin and natamycin, reduced the Vg-prodigiosin uptake by the fungus leading to a significant increase in aflatoxin production, suggesting that uptake is endocytosis-dependent. The Vg treatment also reduced hyphal fusion (>98% inhibition) and branching, which are both endosome-dependent processes. Our results, therefore, collectively support our theory that Vg-associated aflatoxin inhibition is mediated by an endocytosis-dependent uptake of Vg-prodigiosin, which possibly leads to a disruption of normal endosomal functions

The Neuropathology Of Chronic Relapsing Experimental Allergic Encephalomyelitis Induced In The Lewis Rat By Inoculation With Whole Spinal Cord And Treatment With Cyclosporin A

Chronic relapsing experimental allergic encephalomyelitis was induced in Lewis rats by inoculation with guinea-pig spinal cord and complete Freund's adjuvant followed by treatment with low-dose cyclosporin A. In most animals, tail and limb weakness developed in a relapsing remitting pattern but in some these signs were persistent or progressive from onset. Histological studies during the early stages of clinically active disease ( 28 days after inoculation) had extensive spinal cord demyelination but minimal PNS demyelination. In these animals, large plaques of demyelination with gliosis and prominent plasma cells occurred particularly in the thoracic spinal cord, and lesions of different ages were present within the spinal cord. CNS and PNS remyelination with oligodendocytes and Schwann cells respectively was present in all animals studied later than 18 days after inoculation (the time of the first remission, if it occurred). In both early and late clinically active disease electron microscopy revealed macrophages invading and destroying CNS myelin sheaths. Active demyelination was sometimes found in regions of CNS remyelination, suggesting that remyelinated fibres were being attacked. Axonal degeneration occurred in the spinal cord. During clinical remission there was CNS and PNS remyelination and much less inflammation; however active demyelination still occurred to a limited degree

Electroporation-Induced Electrosensitization

BACKGROUND: Electroporation is a method of disrupting the integrity of cell membrane by electric pulses (EPs). Electrical modeling is widely employed to explain and study electroporation, but even most advanced models show limited predictive power. No studies have accounted for the biological consequences of electroporation as a factor that alters the cell's susceptibility to forthcoming EPs. METHODOLOGY/PRINCIPAL FINDINGS: We focused first on the role of EP rate for membrane permeabilization and lethal effects in mammalian cells. The rate was varied from 0.001 to 2,000 Hz while keeping other parameters constant (2 to 3,750 pulses of 60-ns to 9-µs duration, 1.8 to 13.3 kV/cm). The efficiency of all EP treatments was minimal at high rates and started to increase gradually when the rate decreased below a certain value. Although this value ranged widely (0.1-500 Hz), it always corresponded to the overall treatment duration near 10 s. We further found that longer exposures were more efficient irrespective of the EP rate, and that splitting a high-rate EP train in two fractions with 1-5 min delay enhanced the effects severalfold. CONCLUSIONS/SIGNIFICANCE: For varied experimental conditions, EPs triggered a delayed and gradual sensitization to EPs. When a portion of a multi-pulse exposure was delivered to already sensitized cells, the overall effect markedly increased. Because of the sensitization, the lethality in EP-treated cells could be increased from 0 to 90% simply by increasing the exposure duration, or the exposure dose could be reduced twofold without reducing the effect. Many applications of electroporation can benefit from accounting for sensitization, by organizing the exposure either to maximize sensitization (e.g., for sterilization) or, for other applications, to completely or partially avoid it. In particular, harmful side effects of electroporation-based therapies (electrochemotherapy, gene therapies, tumor ablation) include convulsions, pain, heart fibrillation, and thermal damage. Sensitization can potentially be employed to reduce these side effects while preserving or increasing therapeutic efficiency

Evolutionary History of Helicobacter pylori Sequences Reflect Past Human Migrations in Southeast Asia

The human population history in Southeast Asia was shaped by numerous migrations and population expansions. Their reconstruction based on archaeological, linguistic or human genetic data is often hampered by the limited number of informative polymorphisms in classical human genetic markers, such as the hypervariable regions of the mitochondrial DNA. Here, we analyse housekeeping gene sequences of the human stomach bacterium Helicobacter pylori from various countries in Southeast Asia and we provide evidence that H. pylori accompanied at least three ancient human migrations into this area: i) a migration from India introducing hpEurope bacteria into Thailand, Cambodia and Malaysia; ii) a migration of the ancestors of Austro-Asiatic speaking people into Vietnam and Cambodia carrying hspEAsia bacteria; and iii) a migration of the ancestors of the Thai people from Southern China into Thailand carrying H. pylori of population hpAsia2. Moreover, the H. pylori sequences reflect iv) the migrations of Chinese to Thailand and Malaysia within the last 200 years spreading hspEasia strains, and v) migrations of Indians to Malaysia within the last 200 years distributing both hpAsia2 and hpEurope bacteria. The distribution of the bacterial populations seems to strongly influence the incidence of gastric cancer as countries with predominantly hspEAsia isolates exhibit a high incidence of gastric cancer while the incidence is low in countries with a high proportion of hpAsia2 or hpEurope strains. In the future, the host range expansion of hpEurope strains among Asian populations, combined with human motility, may have a significant impact on gastric cancer incidence in Asia

Incidental sounds of locomotion in animal cognition

The highly synchronized formations that characterize schooling in fish and the flight of certain bird groups have frequently been explained as reducing energy expenditure. I present an alternative, or complimentary, hypothesis that synchronization of group movements may improve hearing perception. Although incidental sounds produced as a by-product of locomotion (ISOL) will be an almost constant presence to most animals, the impact on perception and cognition has been little discussed. A consequence of ISOL may be masking of critical sound signals in the surroundings. Birds in flight may generate significant noise; some produce wing beats that are readily heard on the ground at some distance from the source. Synchronization of group movements might reduce auditory masking through periods of relative silence and facilitate auditory grouping processes. Respiratory locomotor coupling and intermittent flight may be other means of reducing masking and improving hearing perception. A distinct border between ISOL and communicative signals is difficult to delineate. ISOL seems to be used by schooling fish as an aid to staying in formation and avoiding collisions. Bird and bat flocks may use ISOL in an analogous way. ISOL and interaction with animal perception, cognition, and synchronized behavior provide an interesting area for future study