Effects of pleiotropic and alleles of on envelope stress and antibiotic sensitivity.

The EnvZ-OmpR two-component system of regulates the expression of the and porin genes in response to medium osmolarity. However, certain mutations in confer pleiotropy by affecting the expression of genes of the iron and maltose regulons not normally controlled by EnvZ-OmpR. In this study, we obtained two novel and pleiotropic alleles, and , among revertants of a mutant with heightened envelope stress and an outer membrane (OM) permeability defect.

Peeling the onion: additional layers of regulation in the acid stress response.

Bacteria are capable of withstanding large changes in osmolality and cytoplasmic pH, unlike eukaryotes that tightly regulate their pH and cellular composition. Previous studies on the bacterial acid stress response described a rapid, brief acidification, followed by immediate recovery. More recent experiments with better pH probes have imaged single living cells, and we now appreciate that following acid stress, bacteria maintain an acidic cytoplasm for as long as the stress remains.

Visualizing the invisible: novel approaches to visualizing bacterial proteins and host-pathogen interactions.

Host-pathogen interactions play a critical role in infectious diseases, and understanding the underlying mechanisms is vital for developing effective therapeutic strategies. The visualization and characterization of bacterial proteins within host cells is key to unraveling the dynamics of these interactions. Various protein labeling strategies have emerged as powerful tools for studying host-pathogen interactions, enabling the tracking, localization, and functional analysis of bacterial proteins in real-time.

RpoS activates formation of Typhi biofilms and drives persistence in the gall bladder.

The development of strategies for targeting the asymptomatic carriage of Typhi in chronic typhoid patients has suffered owing to our basic lack of understanding of the molecular mechanisms that enable the formation of . Typhi biofilms. Traditionally, studies have relied on cholesterol-attached biofilms formed by a closely related serovar, Typhimurium, to mimic multicellular Typhi communities formed on human gallstones.

A pH-sensitive switch activates virulence in .

The transcriptional regulator SsrB acts as a switch between virulent and biofilm lifestyles of non-typhoidal serovar Typhimurium. During infection, phosphorylated SsrB activates genes on Pathogenicity Island-2 (SPI-2) essential for survival and replication within the macrophage. Low pH inside the vacuole is a key inducer of expression and SsrB activation.

Small, Dynamic Clusters of Tir-Intimin Seed Actin Polymerization.

The understanding of actin pedestal formation by enteropathogenic Escherichia coli (EPEC) relies mainly on static ensemble information obtained from cell lysates. Here, the dynamic nature of signaling components on the subsecond timescale, which resemble phase condensates, is demonstrated. Unlike in vitro phase condensates, transfected intimin receptor (Tir) and downstream component form clusters 200 nm in diameter that are spaced ≈500 nm on average, indicating cellular regulation.

The anti-immune dengue subgenomic flaviviral RNA is present in vesicles in mosquito saliva and is associated with increased infectivity.

Mosquito transmission of dengue viruses to humans starts with infection of skin resident cells at the biting site. There is great interest in identifying transmission-enhancing factors in mosquito saliva in order to counteract them. Here we report the discovery of high levels of the anti-immune subgenomic flaviviral RNA (sfRNA) in dengue virus 2-infected mosquito saliva.

Super-Resolution Imaging of Bacterial Secreted Proteins Using Genetic Code Expansion.

Type three secretion systems (T3SSs) enable gram-negative bacteria to inject a battery of effector proteins directly into the cytosol of eukaryotic host cells. Upon entry, the injected effector proteins cooperatively modulate eukaryotic signaling pathways and reprogram cellular functions, enabling bacterial entry and survival. Monitoring and localizing these secreted effector proteins in the context of infections provides a footprint for defining the dynamic interface of host-pathogen interactions.

INTS13 variants causing a recessive developmental ciliopathy disrupt assembly of the Integrator complex.

Oral-facial-digital (OFD) syndromes are a heterogeneous group of congenital disorders characterized by malformations of the face and oral cavity, and digit anomalies. Mutations within 12 cilia-related genes have been identified that cause several types of OFD, suggesting that OFDs constitute a subgroup of developmental ciliopathies. Through homozygosity mapping and exome sequencing of two families with variable OFD type 2, we identified distinct germline variants in INTS13, a subunit of the Integrator complex.

Polyimidazolium Protects against an Invasive Clinical Isolate of Salmonella Typhimurium.

Frequent outbreaks of Salmonella Typhimurium infection, in both animal and human populations and with the potential for zoonotic transmission, pose a significant threat to the public health sector. The rapid emergence and spread of more invasive multidrug-resistant clinical isolates of Salmonella further highlight the need for the development of new drugs with effective broad-spectrum bactericidal activities. The synthesis and evaluation of main-chain cationic polyimidazolium 1 (PIM1) against several Gram-positive and Gram-negative bacteria have previously demonstrated the efficacy profile of PIM1.

Distinct Interaction Mechanism of RNA Polymerase and ResD at Proximal and Distal Subsites for Transcription Activation of Nitrite Reductase in Bacillus subtilis.

The ResD-ResE signal transduction system plays a pivotal role in anaerobic nitrate respiration in Bacillus subtilis. The operon encoding nitrite reductase is essential for nitrate respiration and is tightly controlled by the ResD response regulator. To understand the mechanism of ResD-dependent transcription activation of the operon, we explored ResD-RNA polymerase (RNAP), ResD-DNA, and RNAP-DNA interactions required for transcription.

How Can a Histidine Kinase Respond to Mechanical Stress?

Bacteria respond to physical forces perceived as mechanical stress as part of their comprehensive environmental sensing strategy. Histidine kinases can then funnel diverse environmental stimuli into changes in gene expression through a series of phosphorelay reactions. Because histidine kinases are most often embedded in the inner membrane, they can be sensitive to changes in membrane tension that occurs, for example, in response to osmotic stress, or when deformation of the cell body occurs upon encountering a surface before forming biofilms, or inside the host in response to shear stress in the kidney, intestine, lungs, or blood stream.

Genetic code expansion enables visualization of type three secretion system components and secreted effectors.

Type three secretion systems enable bacterial pathogens to inject effectors into the cytosol of eukaryotic hosts to reprogram cellular functions. It is technically challenging to label effectors and the secretion machinery without disrupting their structure/function. Herein, we present a new approach for labeling and visualization of previously intractable targets.

Super-resolution imaging of bacterial pathogens and visualization of their secreted effectors.

Recent advances in super-resolution imaging techniques, together with new fluorescent probes have enhanced our understanding of bacterial pathogenesis and their interplay within the host. In this review, we provide an overview of what these techniques have taught us about the bacterial lifestyle, the nucleoid organization, its complex protein secretion systems, as well as the secreted virulence factors.

EnvZ/OmpR Two-Component Signaling: An Archetype System That Can Function Noncanonically.

Two-component regulatory systems represent the major paradigm for signal transduction in prokaryotes. The simplest systems are composed of a sensor kinase and a response regulator. The sensor is often a membrane protein that senses a change in environmental conditions and is autophosphorylated by ATP on a histidine residue.

Switching Lifestyles Is an Adaptive Strategy of Bacterial Pathogens.

Gram-positive and Gram-negative pathogens exist as planktonic cells only at limited times during their life cycle. In response to environmental signals such as temperature, pH, osmolality, and nutrient availability, pathogenic bacteria can adopt varied cellular fates, which involves the activation of virulence gene programs and/or the induction of a sessile lifestyle to form multicellular surface-attached communities. In , SsrB is the response regulator which governs the lifestyle switch from an intracellular virulent state to form dormant biofilms in chronically infected hosts.

biofilms program innate immunity for persistence in .

The adaptive in vivo mechanisms underlying the switch in lifestyles from the infectious form to a dormant form remain unknown. We employed as a heterologous host to understand the temporal dynamics of pathogenesis and to identify its lifestyle form in vivo. We discovered that exists as sessile aggregates, or in vivo biofilms, in the persistently infected gut.

Single cell, super-resolution imaging reveals an acid pH-dependent conformational switch in SsrB regulates SPI-2.

After is phagocytosed, it resides in an acidic vacuole. Its cytoplasm acidifies to pH 5.6; acidification activates pathogenicity island 2 (SPI-2).

A New Role of OmpR in Acid and Osmotic Stress in and .

Bacteria survive and respond to diverse environmental conditions and during infection inside the host by systematic regulation of stress response genes. and . Typhimurium can undergo large changes in intracellular osmolality (up to 1.

The role of acid stress in Salmonella pathogenesis.

After uptake by epithelial cells or engulfment by macrophages, Salmonella resides in an acidic vacuole. Salmonella senses this acidic compartment through the action of the EnvZ/OmpR two-component regulatory system. OmpR, in turn, represses the cadC/BA system, preventing neutralization of the bacterial cytoplasm.

Engineering an Osmosensor by Pivotal Histidine Positioning within Disordered Helices.

Histidine kinases (HKs) funnel diverse environmental stimuli into a single autophosphorylation event at a conserved histidine residue. The HK EnvZ is a global sensor of osmolality and cellular acid pH. In previous studies, we discovered that osmosensing in EnvZ was mediated through osmolyte-induced stabilization of the partially disordered helical backbone spanning the conserved histidine autophosphorylation site (His).

A single molecule analysis of H-NS uncouples DNA binding affinity from DNA specificity.

Heat-stable nucleoid structuring protein (H-NS) plays a crucial role in gene silencing within prokaryotic cells and is important in pathogenesis. It was reported that H-NS silences nearly 5% of the genome, yet the molecular mechanism of silencing is not well understood. Here, we employed a highly-sensitive single-molecule counting approach, and measured the dissociation constant (KD) of H-NS binding to single DNA binding sites.

New Variant of Multidrug-Resistant Serovar Typhimurium Associated with Invasive Disease in Immunocompromised Patients in Vietnam.

Nontyphoidal (NTS), particularly serovar Typhimurium, is among the leading etiologic agents of bacterial enterocolitis globally and a well-characterized cause of invasive disease (iNTS) in sub-Saharan Africa. In contrast,  Typhimurium is poorly defined in Southeast Asia, a known hot spot for zoonotic disease with a recently described burden of iNTS disease. Here, we aimed to add insight into the epidemiology and potential impact of zoonotic transfer and antimicrobial resistance (AMR) in  Typhimurium associated with iNTS and enterocolitis in Vietnam.

The Pearling Transition Provides Evidence of Force-Driven Endosomal Tubulation during Infection.

Bacterial pathogens exploit eukaryotic pathways for their own end. Upon ingestion, serovar Typhimurium passes through the stomach and then catalyzes its uptake across the intestinal epithelium. It survives and replicates in an acidic vacuole through the action of virulence factors secreted by a type three secretion system located on pathogenicity island 2 (SPI-2).