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1. Microampere Electric Current Causes Bacterial Membrane Damage and Two-Way Leakage in a Short Period of Time

   https://doi.org/10.1128/AEM.01015-20  

   Krishnamurthi, Venkata Rao ; Rogers, Ariel ; Peifer, Janet ; Niyonshuti, Isabelle I. ; Chen, Jingyi ; Wang, Yong ; Julia Pettinari, M. ( August 2020 , Applied and Environmental Microbiology)

   ABSTRACT Physical agents, such as low electric voltage and current, have recently gained attention for antimicrobial treatment due to their bactericidal capability. Although microampere electric current was shown to suppress the growth of bacteria, it remains unclear to what extent the microampere current damaged the bacterial membrane. Here, we investigated the membrane damage and two-way leakage caused by microampere electric current (≤100 μA) with a short exposure time (30 min). Based on MitoTracker staining, propidium iodide staining, filtration assays, and quantitative single-molecule localization microscopy, we observed significant membrane damage, which allowed two-way leakage of ions, small molecules, and proteins. This study paves the way to new development of antimicrobial applications for ultralow electric voltage and current. IMPORTANCE Although electric voltage and current have been studied for a long time in terms of their ability to suppress the growth of bacteria and to kill bacteria, increasing interest has been aroused more recently due to the prevalence of antibiotic resistance of microbes in past decades. Toward understanding the antimicrobial mechanism of low electric voltage and current, previous studies showed that treating bacteria with milliampere electric currents (≥5 mA) for ≥72 h led to significant damage of the bacterial membrane, which likely resulted in leakage of cellular contents and influx of toxic substances through the damaged membrane. However, it remains unclear to what extent membrane damage and two-way (i.e., inward and outward) leakage are caused by lower (i.e., microampere) electric current in a shorter time frame. In this work, we set out to answer this question. We observed that the membrane damage was caused by microampere electric current in half an hour, which allowed two-way leakage of ions, small molecules, and proteins. 

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2. Silver ions cause oscillation of bacterial length of Escherichia coli

   https://doi.org/10.1038/s41598-019-48113-4  

   Krishnamurthi, Venkata Rao ; Chen, Jingyi ; Wang, Yong ( August 2019 , Scientific Reports)

   Silver (Ag) in various forms have recently gained broad interest and been revisited due to their promising antimicrobial effects. Here we report our study on the morphological dynamics of live bacteria when subjected to Ag⁺ions. Using time-lapse microscopy, we observed oscillations of cell-length for a large fraction of bacteria exposed to 60μM of Ag⁺ions. In addition, we found that the responses of bacteria to Ag⁺ions were heterogeneous. We quantified the oscillations of cell-length with power spectral density, which appeared different from that of bacteria growing in the absence of Ag⁺ions. Furthermore, a model similar to the predator-prey argument was developed to understand the observed oscillations of cell-length upon exposure to Ag⁺ions. This model not only successfully produced the oscillations but also explained the observed heterogeneity in the bacterial responses to Ag⁺ions.

    

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3. Rigidification of the Escherichia coli cytoplasm by the human antimicrobial peptide LL-37 revealed by superresolution fluorescence microscopy

   https://doi.org/10.1073/pnas.1814924116  

   Zhu, Yanyu ; Mohapatra, Sonisilpa ; Weisshaar, James C. ( December 2018 , Proceedings of the National Academy of Sciences)

   Superresolution, single-particle tracking reveals effects of the cationic antimicrobial peptide LL-37 on theEscherichia colicytoplasm. Seconds after LL-37 penetrates the cytoplasmic membrane, the chromosomal DNA becomes rigidified on a length scale of ∼30 nm, evidenced by the loss of jiggling motion of specific DNA markers. The diffusive motion of a subset of ribosomes is also frozen. The mean diffusion coefficients of the DNA-binding protein HU and the nonendogenous protein Kaede decrease twofold. Roughly 10⁸LL-37 copies flood the cell (mean concentration ∼90 mM). Much of the LL-37 remains bound within the cell after extensive rinsing with fresh growth medium. Growth never recovers. The results suggest that the high concentration of adsorbed polycationic peptides forms a dense network of noncovalent, electrostatic linkages within the chromosomal DNA and among 70S-polysomes. The bacterial cytoplasm comprises a concentrated collection of biopolymers that are predominantly polyanionic (e.g., DNA, ribosomes, RNA, and most globular proteins). In normal cells, this provides a kind of electrostatic lubrication, enabling facile diffusion despite high biopolymer volume fraction. However, this same polyanionic nature renders the cytoplasm susceptible to massive adsorption of polycationic agents once penetration of the membranes occurs. If this phenomenon proves widespread across cationic agents and bacterial species, it will help explain why resistance to antimicrobial peptides develops only slowly. The results suggest two design criteria for polycationic peptides that efficiently kill gram-negative bacteria: facile penetration of the outer membrane and the ability to alter the cytoplasm by electrostatically linking double-stranded DNA and 70S-polysomes.

    

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4. Redefining the H-NS protein family: a diversity of specialized core and accessory forms exhibit hierarchical transcriptional network integration

   https://doi.org/10.1093/nar/gkaa709  

   Fitzgerald, Stephen ; Kary, Stefani C ; Alshabib, Ebtihal Y ; MacKenzie, Keith D ; Stoebel, Daniel M ; Chao, Tzu-Chiao ; Cameron, Andrew D ( September 2020 , Nucleic Acids Research)

   null (Ed.)

   Abstract H-NS is a nucleoid structuring protein and global repressor of virulence and horizontally-acquired genes in bacteria. H-NS can interact with itself or with homologous proteins, but protein family diversity and regulatory network overlap remain poorly defined. Here, we present a comprehensive phylogenetic analysis that revealed deep-branching clades, dispelling the presumption that H-NS is the progenitor of varied molecular backups. Each clade is composed exclusively of either chromosome-encoded or plasmid-encoded proteins. On chromosomes, stpA and newly discovered hlpP are core genes in specific genera, whereas hfp and newly discovered hlpC are sporadically distributed. Six clades of H-NS plasmid proteins (Hpp) exhibit ancient and dedicated associations with plasmids, including three clades with fidelity for plasmid incompatibility groups H, F or X. A proliferation of H-NS homologs in Erwiniaceae includes the first observation of potentially co-dependent H-NS forms. Conversely, the observed diversification of oligomerization domains may facilitate stable co-existence of divergent homologs in a genome. Transcriptomic and proteomic analysis in Salmonella revealed regulatory crosstalk and hierarchical control of H-NS homologs. We also discovered that H-NS is both a repressor and activator of Salmonella Pathogenicity Island 1 gene expression, and both regulatory modes are restored by Sfh (HppH) in the absence of H-NS. 

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5. Single‐molecule tracking reveals that the nucleoid‐associated protein HU plays a dual role in maintaining proper nucleoid volume through differential interactions with chromosomal DNA

   https://doi.org/10.1111/mmi.14572  

   Bettridge, Kelsey ; Verma, Subhash ; Weng, Xiaoli ; Adhya, Sankar ; Xiao, Jie ( August 2020 , Molecular Microbiology)

   HU (Histone‐like protein fromEscherichia colistrain U93) is the most conserved nucleoid‐associated protein in eubacteria, but how it impacts global chromosome organization is poorly understood. Using single‐molecule tracking, we demonstrate that HU exhibits nonspecific, weak, and transitory interactions with the chromosomal DNA. These interactions are largely mediated by three conserved, surface‐exposed lysine residues (triK), which were previously shown to be responsible for nonspecific binding to DNA. The loss of these weak, transitory interactions in a HUα(triKA) mutant results in an over‐condensed and mis‐segregated nucleoid. Mutating a conserved proline residue (P63A) in the HUα subunit, deleting the HUβ subunit, or deleting nucleoid‐associated naRNAs, each previously implicated in HU’s high‐affinity binding to kinked or cruciform DNA, leads to less dramatically altered interacting dynamics of HU compared to the HUα(triKA) mutant, but highly expanded nucleoids. Our results suggest HU plays a dual role in maintaining proper nucleoid volume through its differential interactions with chromosomal DNA. On the one hand, HU compacts the nucleoid through specific DNA structure‐binding interactions. On the other hand, it decondenses the nucleoid through many nonspecific, weak, and transitory interactions with the bulk chromosome. Such dynamic interactions may contribute to the viscoelastic properties and fluidity of the bacterial nucleoid to facilitate proper chromosome functions.

    

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