ABSTRACT
Bacterial cells frequently experience fluid motion in their natural environments, like water bodies, aerosols, fomites, human capillaries, etc., a phenomenon that researchers have largely overlooked. Nevertheless, some reports have suggested that the interfacial stresses caused by fluid motion inside evaporating droplets or shear flows within capillaries may trigger physiological and morphological changes in the bacterial cells. Remarkably, the virulence of bacterial cells exhibits significant alterations in response to fluctuations in stress levels and external environmental factors. The dynamics of bacterial systems are analogous to colloidal systems but with the distinction that bacterial systems exhibit responsiveness, necessitating thorough exploration in dynamic environments. In this perspective, we discuss the important issue pertaining to bacterial survival, virulence, and disease transmission. Furthermore, we delineate a pathway and underscore emerging opportunities that demand exploration to unveil new avenues in the domains of bacterial pathogenicity, drug development, and strategies for disease mitigation.
ABSTRACT
HYPOTHESIS: The study shows for the first time a fivefold difference in the survivability of the bacterium Pseudomonas Aeruginosa (PA) in a realistic respiratory fluid droplet on fomites undergoing drying at different environmental conditions. For instance, in 2023, the annual average outdoor relative humidity (RH) and temperature in London (UK) is 71 % and 11 °C, whereas in New Delhi (India), it is 45 % and 26 °C, showing that disease spread from fomites could have a demographic dependence. Respiratory fluid droplet ejections containing pathogens on inanimate surfaces are crucial in disease spread, especially in nosocomial settings. However, the interplay between evaporation dynamics, internal fluid flow and precipitation and their collective influence on the distribution and survivability of pathogens at different environmental conditions are less known. EXPERIMENTS: Shadowgraphy imaging is employed to study evaporation, and optical microscopy imaging is used for precipitation dynamics. Micro-particle image velocimetry (MicroPIV) measurements reveal the internal flow dynamics. Confocal imaging of fluorescently labelled PA elucidates the bacterial distribution within the deposits. FINDINGS: The study finds that the evaporation rate is drastically impeded during drying at elevated solutal concentrations, particularly at high RH and low temperature conditions. MicroPIV shows reduced internal flow under high RH and low temperature (low evaporation rate) conditions. Evaporation rate influences crystal growth, with delayed efflorescence and extending crystallization times. PA forms denser peripheral arrangements under high evaporation rates and shows a fivefold increase in survivability under low evaporation rates. These findings highlight the critical impact of environmental conditions on pathogen persistence and disease spread from inanimate surfaces.
Subject(s)
Pseudomonas aeruginosa , Surface Properties , Pseudomonas aeruginosa/isolation & purification , Weather , Humidity , Particle Size , Microbial Viability , Fomites/microbiology , DesiccationABSTRACT
Our previous study showed that OmpA-deficient Salmonella Typhimurium (STM) failed to retain LAMP-1, quit Salmonella-containing vacuole (SCV) and escaped to the host cytosol. Here we show that the cytosolic population of STM ΔompA sequestered autophagic markers, syntaxin17 and LC3B in a sseL-dependent manner and initiated lysosomal fusion. Moreover, inhibition of autophagy using bafilomycinA1 restored its intracellular proliferation. Ectopic overexpression of OmpA in STM ΔsifA restored its vacuolar niche and increased interaction of LAMP-1, suggesting a sifA-independent role of OmpA in maintaining an intact SCV. The OmpA extracellular loops impaired the LAMP-1 recruitment to SCV and caused bacterial release into the cytosol of macrophages, but unlike STM ΔompA, they retained their outer membrane stability and didn't activate the lysosomal degradation pathway aiding in their intra-macrophage survival. Finally, OmpA extracellular loop mutations protected the cytosolic STM ΔsifA from the lysosomal surveillance, revealing a unique OmpA-dependent strategy of STM for its intracellular survival.
ABSTRACT
Salmonella infection entails a cascade of attacks and defence measures. After breaching the intestinal epithelial barrier, Salmonella is phagocytosed by macrophages, where the bacteria encounter multiple stresses, to which it employs relevant countermeasures. Our study shows that, in Salmonella, the polyamine spermidine activates a stress response mechanism by regulating critical antioxidant genes. Salmonella Typhimurium mutants for spermidine transport and synthesis cannot mount an antioxidative response, resulting in high intracellular ROS levels. These mutants are also compromised in their ability to be phagocytosed by macrophages. Furthermore, it regulates a novel enzyme in Salmonella, Glutathionyl-spermidine synthetase (GspSA), which prevents the oxidation of proteins in E. coli. Moreover, the spermidine mutants and the GspSA mutant show significantly reduced survival in the presence of hydrogen peroxide in vitro and reduced organ burden in the mouse model of Salmonella infection. Conversely, in macrophages isolated from gp91phox-/- mice, we observed a rescue in the attenuated fold proliferation previously observed upon infection. We found that Salmonella upregulates polyamine biosynthesis in the host through its effectors from SPI-1 and SPI-2, which addresses the attenuated proliferation observed in spermidine transport mutants. Thus, inhibition of this pathway in the host abrogates the proliferation of Salmonella Typhimurium in macrophages. From a therapeutic perspective, inhibiting host polyamine biosynthesis using an FDA-approved chemopreventive drug, D, L-α-difluoromethylornithine (DFMO), reduces Salmonella colonisation and tissue damage in the mouse model of infection while enhancing the survival of infected mice. Therefore, our work provides a mechanistic insight into the critical role of spermidine in stress resistance of Salmonella. It also reveals a bacterial strategy in modulating host metabolism to promote their intracellular survival and shows the potential of DFMO to curb Salmonella infection.
Subject(s)
Bacterial Proteins , Macrophages , Membrane Proteins , NADPH Oxidase 2 , Reactive Oxygen Species , Salmonella typhimurium , Spermidine , Animals , Salmonella typhimurium/metabolism , Salmonella typhimurium/drug effects , Spermidine/metabolism , Mice , Macrophages/microbiology , Macrophages/metabolism , Macrophages/drug effects , Reactive Oxygen Species/metabolism , Bacterial Proteins/metabolism , Bacterial Proteins/genetics , Polyamines/metabolism , Phagocytosis/drug effects , Salmonella Infections/microbiology , Salmonella Infections/metabolism , NADPH Oxidases/metabolism , NADPH Oxidases/genetics , Host-Pathogen Interactions , Spermidine Synthase/metabolism , Spermidine Synthase/genetics , Oxidative Stress/drug effectsABSTRACT
Gut microbiota plays several roles in the host organism's metabolism and physiology. This phenomenon holds across different species from different kingdoms and classes. Different species across various classes engage in continuous crosstalk via various mechanisms with their gut microbiota, ensuring homeostasis of the host. In this Review, the diversity of the microflora, the development of the microflora in the host, its regulations by the host, and its functional implications on the host, especially in the context of dysbiosis, are discussed across different organisms from sponges to humans. Overall, our review aims to address the indispensable nature of the microbiome in the host's survival, fitness, and protection against invading pathogens.
Subject(s)
Gastrointestinal Microbiome , Microbiota , HumansABSTRACT
Spermidine is a poly-cationic molecule belonging to the family of polyamines and is ubiquitously present in all organisms. Salmonella synthesizes, and harbours specialized transporters to import spermidine. A group of polyamines have been shown to assist in Salmonella Typhimurium's virulence and regulation of Salmonella pathogenicity Inslad 1 (SPI-1) genes and stress resistance; however, the mechanism remains elusive. The virulence trait of Salmonella depends on its ability to employ multiple surface structures to attach and adhere to the surface of the target cells before invasion and colonization of the host niche. Our study discovers the mechanism by which spermidine assists in the early stages of Salmonella pathogenesis. For the first time, we report that Salmonella Typhimurium regulates spermidine transport and biosynthesis processes in a mutually inclusive manner. Using a mouse model, we show that spermidine is critical for invasion into the murine Peyer's patches, which further validated our in vitro cell line observation. We show that spermidine controls the mRNA expression of fimbrial (fimA) and non-fimbrial adhesins (siiE, pagN) in Salmonella and thereby assists in attachment to host cell surfaces. Spermidine also regulated the motility through the expression of flagellin genes by enhancing the translation of sigma-28, which features an unusual start codon and a poor Shine-Dalgarno sequence. Besides regulating the formation of the adhesive structures, spermidine tunes the expression of the two-component system BarA/SirA to regulate SPI-1 encoded genes. Thus, our study unravels a novel regulatory mechanism by which spermidine exerts critical functions during Salmonella Typhimurium pathogenesis.
Subject(s)
Salmonella typhimurium , Spermidine , Animals , Mice , Salmonella typhimurium/metabolism , Spermidine/metabolism , Bacterial Proteins/genetics , Bacterial Proteins/metabolism , Flagellin/genetics , Polyamines/metabolism , Gene Expression Regulation, BacterialABSTRACT
Docosahexaenoic acid (DHA) is known to have numerous health benefits and immense dietary value. There is a pressing need to have a deeper understanding of DHA metabolism. Acyl CoA: Diacylglycerol Acyltransferase (DGAT) is an important enzyme of lipid anabolism and an essential piece of the puzzle. Aurantiochytrium limacinum, a primary producer of DHA, is a good model for studying DHA metabolism. Thus, we aimed to investigate important lipid metabolic genes from A. limacinum. We cloned four putative DGATs (DGAT2a, DGAT2b, DGAT2c, and DGAT2d) from A. limacinum and performed detailed in vivo and in vitro characterization. Functional characterization showed that not all the studied genes exhibited DGAT activity. DGAT2a and DGAT2d conferred DGAT activity whereas DGAT2b showed wax synthase (WS) activity and DGAT2c showed dual function of both WS and DGAT. Based on their identified function, DGAT2b and DGAT2c were renamed as AlWS and AlWS/DGAT respectively. DGAT2a was found to exhibit a preference for DHA as a substrate. DGAT2d was found to have robust activity and emerged as a promising candidate for genetic engineering aimed at increasing oil yield. The study enriches our knowledge of lipid biosynthetic enzymes in A. limacinum, which can be utilized to design suitable application strategies.
Subject(s)
Diacylglycerol O-Acyltransferase , Genetic Engineering , Diacylglycerol O-Acyltransferase/genetics , Diacylglycerol O-Acyltransferase/metabolism , LipidsABSTRACT
Immune cells employ diverse mechanisms for host defense. Macrophages, in response to TLR activation, assemble aggresome-like induced structures (ALIS). Our group has shown TLR4-signaling transcriptionally upregulates p62/sequestome1, which assembles ALIS. We have demonstrated that TLR4-mediated autophagy is, in fact, selective-autophagy of ALIS. We hypothesize that TLR-mediated autophagy and ALIS contribute to host-defense. Here we show that ALIS are assembled in macrophages upon exposure to different bacteria. These structures are associated with pathogen-containing phagosomes. Importantly, we present evidence of increased bacterial burden, where ALIS assembly is prevented with p62-specific siRNA. We have employed 3D-super-resolution structured illumination microscopy (3D-SR-SIM) and mass-spectrometric (MS) analyses to gain insight into the assembly of ALIS. Ultra-structural analyses of known constituents of ALIS (p62, ubiquitin, LC3) reveal that ALIS are organized structures with distinct patterns of alignment. Furthermore, MS-analyses of ALIS identified, among others, several proteins of known antimicrobial properties. We have validated MS data by testing the association of some of these molecules (Bst2, IFITM2, IFITM3) with ALIS and the phagocytosed-bacteria. We surmise that AMPs enrichment in ALIS leads to their delivery to bacteria-containing phagosomes and restricts the bacteria. Our findings in this paper support hitherto unknown functions of ALIS in host-defense.
Subject(s)
Antimicrobial Peptides , Toll-Like Receptor 4 , Toll-Like Receptor 4/genetics , Macrophages/metabolism , Ubiquitin/metabolism , Autophagy/physiologyABSTRACT
Salmonella Typhi, the invasive serovar of S. enterica subspecies enterica, causes typhoid fever in healthy human hosts. The emergence of antibiotic-resistant strains has consistently challenged the successful treatment of typhoid fever with conventional antibiotics. Antimicrobial resistance (AMR) in Salmonella is acquired either by mutations in the genomic DNA or by acquiring extrachromosomal DNA via horizontal gene transfer. In addition, Salmonella can form a subpopulation of antibiotic persistent (AP) cells that can survive at high concentrations of antibiotics. These have reduced the effectiveness of the first and second lines of antibiotics used to treat Salmonella infection. The recurrent and chronic carriage of S. Typhi in human hosts further complicates the treatment process, as a remarkable shift in the immune response from pro-inflammatory Th1 to anti-inflammatory Th2 is observed. Recent studies have also highlighted the overlap between AP, persistent infection (PI) and AMR. These incidents have revealed several areas of research. In this review, we have put forward a timeline for the evolution of antibiotic resistance in Salmonella and discussed the different mechanisms of the same availed by the pathogen at the genotypic and phenotypic levels. Further, we have presented a detailed discussion on Salmonella antibiotic persistence (AP), PI, the host and bacterial virulence factors that can influence PI, and how both AP and PI can lead to AMR.
Subject(s)
Salmonella Infections , Typhoid Fever , Humans , Salmonella typhi/genetics , Typhoid Fever/drug therapy , Typhoid Fever/microbiology , Anti-Bacterial Agents/pharmacology , Salmonella Infections/drug therapy , DNA , Microbial Sensitivity TestsABSTRACT
In the tug-of-war between host and pathogen, both evolve to combat each other's defence arsenals. Intracellular phagosomal bacteria have developed strategies to modify the vacuolar niche to suit their requirements best. Conversely, the host tries to target the pathogen-containing vacuoles towards the degradative pathways. The host cells use a robust system through intracellular trafficking to maintain homeostasis inside the cellular milieu. In parallel, intracellular bacterial pathogens have coevolved with the host to harbour strategies to manipulate cellular pathways, organelles, and cargoes, facilitating the conversion of the phagosome into a modified pathogen-containing vacuole (PCV). Key molecular regulators of intracellular traffic, such as changes in the organelle (phospholipid) composition, recruitment of small GTPases and associated effectors, soluble N-ethylmaleimide-sensitive factor-activating protein receptors (SNAREs), etc., are hijacked to evade lysosomal degradation. Legionella, Salmonella, Coxiella, Chlamydia, Mycobacterium, and Brucella are examples of pathogens which diverge from the endocytic pathway by using effector-mediated mechanisms to overcome the challenges and establish their intracellular niches. These pathogens extensively utilise and modulate the end processes of secretory pathways, particularly SNAREs, in repurposing the PCV into specialised compartments resembling the host organelles within the secretory network; at the same time, they avoid being degraded by the host's cellular mechanisms. Here, we discuss the recent research advances on the host-pathogen interaction/crosstalk that involves host SNAREs, conserved cellular processes, and the ongoing host-pathogen defence mechanisms in the molecular arms race against each other. The current knowledge of SNAREs, and intravacuolar bacterial pathogen interactions, enables us to understand host cellular innate immune pathways, maintenance of homeostasis, and potential therapeutic strategies to combat ever-growing antimicrobial resistance.
ABSTRACT
In the present work, experiments are conducted to understand the consequence of stresses generated by flowing fluid on the bacterial morphology and virulence in microfluidic channels. We consider Klebsiella pneumoniae (KP, a clinical isolate), an ESKAPE pathogen, to be the model bacteria responsible for blood stream infections, bacteremia, including pneumonia, urinary tract infections and more. Four different stress conditions are generated by changing the flow rate and channel geometry subsequently altering the shear rate and stressing time (τ). We observe significant changes in the structural aspects of the stressed bacteria. With an increase in stressing parameters, the viability of the bacterial sample deteriorated. Most importantly, these stressed samples proliferate much more than unstressed samples inside the RAW264.7 murine macrophages. The results shed light on the complex relationship between flow stresses and bacterial virulence. Furthermore, the bacterial samples are challenged with ciprofloxacin to see how they behave under different stress conditions. The observations presented in the present study can be extended to model deadly diseases including bacteremia using organ-on-a-chip technology and to understand bacterial pathogenicity under realistic environments.
Subject(s)
Bacteremia , Klebsiella Infections , Animals , Mice , Klebsiella pneumoniae , Klebsiella Infections/drug therapy , Klebsiella Infections/microbiology , Bacteremia/drug therapy , Bacteremia/microbiology , Anti-Bacterial AgentsABSTRACT
Invasive-Non-Typhoidal Salmonella (iNTS) are the major cause of health concern in the low-income, under-developed nations in Africa and Asia that lack proper sanitation facilities. Around 5% of the NTS cases give rise to invasive, extraintestinal diseases leading to focal infections like osteomyelitis, meningitis, osteoarthritis, endocarditis and neonatal sepsis. iNTS serovars like S. Typhimurium, S. Enteritidis, S. Dublin, S. Choleraesuis show a greater propensity to become invasive than others which hints at the genetic basis of their emergence. The major risk factors attributing to the invasive diseases include immune-compromised individuals having co-infection with malaria or HIV, or suffering from malnutrition. The rampant use of antibiotics leading to the emergence of multi-drug resistant strains poses a great challenge in disease management. An extensive understanding of the iNTS pathogenesis and its epidemiology will open up avenues for the development of new vaccination and therapeutic strategies to restrict the spread of this neglected disease.
Subject(s)
Salmonella Infections , Typhoid Fever , Infant, Newborn , Humans , Salmonella Infections/epidemiology , Salmonella/genetics , Anti-Bacterial Agents/pharmacology , AfricaABSTRACT
Host immune response to COVID-19 plays a significant role in regulating disease severity. Although big data analysis has provided significant insights into the host biology of COVID-19 across the world, very few such studies have been performed in the Indian population. This study utilizes a transcriptome-integrated network analysis approach to compare the immune responses between asymptomatic or mild and moderate-severe COVID-19 patients in an Indian cohort. An immune suppression phenotype is observed in the early stages of moderate-severe COVID-19 manifestation. A number of pathways are identified that play crucial roles in the host control of the disease such as the type I interferon response and classical complement pathway which show different activity levels across the severity spectrum. This study also identifies two transcription factors, IRF7 and ESR1, to be important in regulating the severity of COVID-19. Overall this study provides a deep understanding of the peripheral immune landscape in the COVID-19 severity spectrum in the Indian genetic background and opens up future research avenues to compare immune responses across global populations.
Subject(s)
COVID-19 , Interferon Type I , Humans , COVID-19/genetics , Gene Expression Profiling , Phenotype , Transcription FactorsABSTRACT
Salmonella Typhi (S. Typhi), the invasive typhoidal serovar of Salmonella enterica that causes typhoid fever in humans, is a severe threat to global health. It is one of the major causes of high morbidity and mortality in developing countries. According to recent WHO estimates, approximately 11-21 million typhoid fever illnesses occur annually worldwide, accounting for 0.12-0.16 million deaths. Salmonella infection can spread to healthy individuals by the consumption of contaminated food and water. Typhoid fever in humans sometimes is accompanied by several other critical extraintestinal complications related to the central nervous system, cardiovascular system, pulmonary system, and hepatobiliary system. Salmonella Pathogenicity Island-1 and Salmonella Pathogenicity Island-2 are the two genomic segments containing genes encoding virulent factors that regulate its invasion and systemic pathogenesis. This Review aims to shed light on a comparative analysis of the virulence and pathogenesis of the typhoidal and nontyphoidal serovars of S. enterica.
ABSTRACT
Salmonella is a genus of widely spread Gram negative, facultative anaerobic bacteria, which is known to cause »th of diarrheal morbidity and mortality globally. It causes typhoid fever and gastroenteritis by gaining access to the host gut through contaminated food and water. Salmonella utilizes its biofilm lifestyle to strongly resist antibiotics and persist in the host. Although biofilm removal or dispersal has been studied widely, the inhibition of the initiation of Salmonella Typhimurium (STM WT) biofilm remains elusive. This study demonstrates the anti-biofilm property of the cell-free supernatant obtained from a carbon-starvation induced proline peptide transporter mutant (STM ΔyjiY) strain. The STM ΔyjiY culture supernatant primarily inhibits biofilm initiation by regulating biofilm-associated transcriptional network that is reversed upon complementation (STM ΔyjiY:yjiY). We demonstrate that abundance of FlgM correlates with the absence of flagella in the STM ΔyjiY supernatant treated WT cells. NusG works synergistically with the global transcriptional regulator H-NS. Relatively low abundances of flavoredoxin, glutaredoxin, and thiol peroxidase might lead to accumulation of ROS within the biofilm, and subsequent toxicity in STM ΔyjiY supernatant. This work further suggests that targeting these oxidative stress relieving proteins might be a good choice to reduce Salmonella biofilm.
Subject(s)
Salmonella typhimurium , Typhoid Fever , Humans , Salmonella typhimurium/genetics , Salmonella typhimurium/metabolism , Membrane Transport Proteins/metabolism , Biofilms , Proline/metabolismABSTRACT
Despite extensive studies on kinematic features of impacting drops, the effect of mechanical stress on desiccated bacteria-laden droplets remains unexplored. In the present study, we unveiled the consequences of the impaction of bacteria-laden droplets on solid surfaces and their subsequent desiccation on the virulence of an enteropathogen Salmonella typhimurium (STM). The methodology elucidated the deformation, cell-cell interactions, adhesion energy, and roughness in bacteria induced by impact velocity and low moisture because of evaporation. Salmonella retrieved from the dried droplets were used to understand fomite-mediated pathogenesis. The impact velocity-induced mechanical stress deteriorated the in vitro viability of Salmonella. Of interest, an uninterrupted bacterial proliferation was observed in macrophages at higher mechanical stress. Wild-type Salmonella under mechanical stress induced the expression of phoP whereas infecting macrophages. The inability of STM ΔphoP to grow in nutrient-rich dried droplets signifies the role of phoP in sensing the mechanical stress and maintaining the virulence of Salmonella.
ABSTRACT
The bacterial toxin CcdB (Controller of Cell death or division B) targets DNA Gyrase, an essential bacterial topoisomerase, which is also the molecular target for fluoroquinolones. Here, we present a short cell-penetrating 24-mer peptide, CP1-WT, derived from the Gyrase-binding region of CcdB and examine its effect on growth of Escherichia coli, Salmonella Typhimurium, Staphylococcus aureus and a carbapenem- and tigecycline-resistant strain of Acinetobacter baumannii in both axenic cultures and mouse models of infection. The CP1-WT peptide shows significant improvement over ciprofloxacin in terms of its in vivo therapeutic efficacy in treating established infections of S. Typhimurium, S. aureus and A. baumannii. The molecular mechanism likely involves inhibition of Gyrase or Topoisomerase IV, depending on the strain used. The study validates the CcdB binding site on bacterial DNA Gyrase as a viable and alternative target to the fluoroquinolone binding site.
Subject(s)
Anti-Bacterial Agents , Staphylococcus aureus , Animals , Mice , Staphylococcus aureus/genetics , Staphylococcus aureus/metabolism , Anti-Bacterial Agents/pharmacology , DNA Gyrase/chemistry , DNA Gyrase/genetics , DNA Gyrase/metabolism , DNA Topoisomerase IV/genetics , DNA Topoisomerase IV/metabolism , DNA Topoisomerase IV/pharmacology , Peptides/pharmacologyABSTRACT
Rheumatic heart disease (RHD) continues to affect developing countries with low income due to the lack of resources and effective diagnostic techniques. Understanding the genetic basis common to both the diseases and that of progression from its prequel disease state, Acute Rheumatic Fever (ARF), would aid in developing predictive biomarkers and improving patient care. To gain system-wide molecular insights into possible causes for progression, in this pilot study, we collected blood transcriptomes from ARF (5) and RHD (5) patients. Using an integrated transcriptome and network analysis approach, we identified a subnetwork comprising the most significantly differentially expressed genes and most perturbed pathways in RHD compared to ARF. For example, the chemokine signaling pathway was seen to be upregulated, while tryptophan metabolism was found to be downregulated in RHD. The subnetworks of variation between the two conditions provide unbiased molecular-level insights into the host processes that may be linked with the progression of ARF to RHD, which has the potential to inform future diagnostics and therapeutic strategies. We also found a significantly raised neutrophil/lymphocyte ratio in both ARF and RHD cohorts. Activated neutrophils and inhibited Natural Killer cell gene signatures reflected the drivers of the inflammatory process typical to both disease conditions.
Subject(s)
Rheumatic Fever , Rheumatic Heart Disease , Humans , Rheumatic Fever/genetics , Rheumatic Heart Disease/genetics , Rheumatic Heart Disease/diagnosis , Pilot Projects , PovertyABSTRACT
Salmonella, a stealthy facultative intracellular pathogen, utilises an array of host immune evasion strategies. This facilitates successful survival via replicative niche establishment in otherwise hostile environments such as macrophages. Salmonella survives in and utilises macrophages for effective dissemination, ultimately leading to systemic infection. Bacterial xenophagy or macro-autophagy is an important host defense mechanism in macrophages. Here, we report for the first time that the Salmonella pathogenicity island-1 (SPI-1) effector SopB is involved in subverting host autophagy via dual mechanisms. SopB is a phosphoinositide phosphatase capable of altering the phosphoinositide dynamics of the host cell. Here, we demonstrate that SopB mediates escape from autophagy by inhibiting the terminal fusion of Salmonella-containing vacuoles (SCVs) with lysosomes and/or autophagosomes. We also report that SopB downregulates overall lysosomal biogenesis by modulating the Akt-transcription factor EB (TFEB) axis via restricting the latter's nuclear localisation. TFEB is a master regulator of lysosomal biogenesis and autophagy. This reduces the overall lysosome content inside host macrophages, further facilitating the survival of Salmonella in macrophages and systemic dissemination of Salmonella.
Subject(s)
Macroautophagy , Salmonella typhimurium , Autophagy , Bacterial Proteins , Macrophages/microbiology , Salmonella typhimurium/physiologyABSTRACT
Intracellular membrane fusion is mediated by membrane-bridging complexes of soluble N-ethylmaleimide-sensitive factor attachment protein receptors (SNAREs). SNARE proteins are one of the key players in vesicular transport. Several reports shed light on intracellular bacteria modulating host SNARE machinery to establish infection successfully. The critical SNAREs in macrophages responsible for phagosome maturation are Syntaxin 3 (STX3) and Syntaxin 4 (STX4). Reports also suggest that Salmonella actively modulates its vacuole membrane composition to escape lysosomal fusion. Salmonella containing vacuole (SCV) harbours recycling endosomal SNARE Syntaxin 12 (STX12). However, the role of host SNAREs in SCV biogenesis and pathogenesis remains unclear. Upon knockdown of STX3, we observed a reduction in bacterial proliferation, which is concomitantly restored upon the overexpression of STX3. Live-cell imaging of Salmonella-infected cells showed that STX3 localises to the SCV membranes and thus might help in the fusion of SCV with intracellular vesicles to acquire membrane for its division. We also found the interaction STX3-SCV was abrogated when we infected with SPI-2 encoded Type 3 secretion system (T3SS) apparatus mutant (STM ∆ssaV) but not with SPI-1 encoded T3SS apparatus mutant (STM ∆invC). These observations were also consistent in the mice model of Salmonella infection. Together, these results shed light on the effector molecules secreted through T3SS encoded by SPI-2, possibly involved in interaction with host SNARE STX3, which is essential to maintain the division of Salmonella in SCV and help to maintain a single bacterium per vacuole.