Oh my gut! Is the microbial origin of neurodegenerative diseases real?

There is no cure or effective treatment for neurodegenerative protein conformational diseases (PCDs), such as Alzheimer's or Parkinson's diseases, mainly because the etiology of these diseases remains elusive. Recent data suggest that unique changes in the gut microbial composition are associated with these ailments; however, our current understanding of the bacterial role in the pathogenesis of PCDs is hindered by the complexity of the microbial communities associated with specific microbiomes, such as the gut, oral, or vaginal microbiota. The composition of these specific microbiomes is regarded as a unique fingerprint affected by factors such as infections, diet, lifestyle, and antibiotics. All of these factors also affect the severity of neurodegenerative diseases. The majority of studies that reveal microbial contribution are correlational, and various models, including worm, fly, and mouse, are being utilized to decipher the role of individual microbes that may affect disease onset and progression. Recent evidence from across model organisms and humans shows a positive correlation between the presence of gram-negative enteropathogenic bacteria and the pathogenesis of PCDs. While these correlational studies do not provide a mechanistic explanation, they do reveal contributing bacterial species and provide an important basis for further investigation. One of the lurking concerns related to the microbial contribution to PCDs is the increasing prevalence of antibiotic resistance and poor antibiotic stewardship, which ultimately select for proteotoxic bacteria, especially the gram-negative species that are known for intrinsic resistance. In this review, we summarize what is known about individual microbial contribution to PCDs and the potential impact of increasing antimicrobial resistance.

Colonization of the Caenorhabditis elegans gut with human enteric bacterial pathogens leads to proteostasis disruption that is rescued by butyrate.

Protein conformational diseases are characterized by misfolding and toxic aggregation of metastable proteins, often culminating in neurodegeneration. Enteric bacteria influence the pathogenesis of neurodegenerative diseases; however, the complexity of the human microbiome hinders our understanding of how individual microbes influence these diseases. Disruption of host protein homeostasis, or proteostasis, affects the onset and progression of these diseases. To investigate the effect of bacteria on host proteostasis, we used Caenorhabditis elegans expressing tissue-specific polyglutamine reporters that detect changes in the protein folding environment. We found that colonization of the C. elegans gut with enteric bacterial pathogens disrupted proteostasis in the intestine, muscle, neurons, and the gonad, while the presence of bacteria that conditionally synthesize butyrate, a molecule previously shown to be beneficial in neurodegenerative disease models, suppressed aggregation and the associated proteotoxicity. Co-colonization with this butyrogenic strain suppressed bacteria-induced protein aggregation, emphasizing the importance of microbial interaction and its impact on host proteostasis. Further experiments demonstrated that the beneficial effect of butyrate depended on the bacteria that colonized the gut and that this protective effect required SKN-1/Nrf2 and DAF-16/FOXO transcription factors. We also found that bacteria-derived protein aggregates contribute to the observed disruption of host proteostasis. Together, these results reveal the significance of enteric infection and gut dysbiosis on the pathogenesis of protein conformational diseases and demonstrate the potential of using butyrate-producing microbes as a preventative and treatment strategy for neurodegenerative disease.

Bacteria-Derived Protein Aggregates Contribute to the Disruption of Host Proteostasis.

Neurodegenerative protein conformational diseases are characterized by the misfolding and aggregation of metastable proteins encoded within the host genome. The host is also home to thousands of proteins encoded within exogenous genomes harbored by bacteria, fungi, and viruses. Yet, their contributions to host protein-folding homeostasis, or proteostasis, remain elusive. Recent studies, including our previous work, suggest that bacterial products contribute to the toxic aggregation of endogenous host proteins. We refer to these products as bacteria-derived protein aggregates (BDPAs). Furthermore, antibiotics were recently associated with an increased risk for neurodegenerative diseases, including Parkinson's disease and amyotrophic lateral sclerosis-possibly by virtue of altering the composition of the human gut microbiota. Other studies have shown a negative correlation between disease progression and antibiotic administration, supporting their protective effect against neurodegenerative diseases. These contradicting studies emphasize the complexity of the human gut microbiota, the gut-brain axis, and the effect of antibiotics. Here, we further our understanding of bacteria's effect on host protein folding using the model Caenorhabditis elegans. We employed genetic and chemical methods to demonstrate that the proteotoxic effect of bacteria on host protein folding correlates with the presence of BDPAs. Furthermore, the abundance and proteotoxicity of BDPAs are influenced by gentamicin, an aminoglycoside antibiotic that induces protein misfolding, and by butyrate, a short-chain fatty acid that we previously found to affect host protein aggregation and the associated toxicity. Collectively, these results increase our understanding of host-bacteria interactions in the context of protein conformational diseases.

Antibiotic prescribing patterns for acute respiratory infections in a free clinic network: A pooled cross-sectional study.

In the United States, overprescribing of antibiotics for viral respiratory infections and antimicrobial resistance continue to be public health concerns. To date, no literature has focused on antibiotic prescribing patterns from free clinics. To address this gap, we used patient-level data from a student-run free clinic network of four primary care clinics to assess factors associated with inappropriate antibiotic prescribing for viral respiratory infections. Treatment plans were deemed inappropriate if any type of antibiotic was prescribed. We used unpaired t-tests and chi-square tests to assess for differences in receiving an inappropriate antibiotic prescription by patient-level factors (i.e., age, race/ethnicity, sex, educational attainment, preferred language, insurance status). Of 298 visits, 22.5% did not meet treatment guidelines. No patient-level factors studied were associated with inappropriate antibiotic prescribing. Our findings suggest other factors, beyond patient-level, may be drivers of variation in antibiotic prescribing in free clinics.

GigC, a LysR Family Transcription Regulator, Is Required for Cysteine Metabolism and Virulence in Acinetobacter baumannii.

A critical facet of mammalian innate immunity involves the hosts' attempts to sequester and/or limit the availability of key metabolic products from pathogens. For example, nutritional immunity encompasses host approaches to limit the availability of key heavy metal ions such as zinc and iron. Previously, we identified several hundred genes in a multidrug-resistant isolate of Acinetobacter baumannii that are required for growth and/or survival in the Galleria mellonella infection model. In the present study, we further characterize one of these genes, a LysR family transcription regulator that we previously named GigC. We show that mutant strains lacking gigC have impaired growth in the absence of the amino acid cysteine and that gigC regulates the expression of several genes involved in the sulfur assimilation and cysteine biosynthetic pathways. We further show that cells harboring a deletion of the gigC gene are attenuated in two murine infection models, suggesting that the GigC protein, likely through its regulation of the cysteine biosynthetic pathway, plays a key role in the virulence of A. baumannii.

Periplasmic protein EipA determines envelope stress resistance and virulence in Brucella abortus.

Molecular components of the Brucella abortus cell envelope play a major role in its ability to infect, colonize and survive inside mammalian host cells. In this study, we have defined a role for a conserved gene of unknown function in B. abortus envelope stress resistance and infection. Expression of this gene, which we name eipA, is directly activated by the essential cell cycle regulator, CtrA. eipA encodes a soluble periplasmic protein that adopts an unusual eight-stranded ß-barrel fold. Deletion of eipA attenuates replication and survival in macrophage and mouse infection models, and results in sensitivity to treatments that compromise the cell envelope integrity. Transposon disruption of genes required for LPS O-polysaccharide biosynthesis is synthetically lethal with eipA deletion. This genetic connection between O-polysaccharide and eipA is corroborated by our discovery that eipA is essential in Brucella ovis, a naturally rough species that harbors mutations in several genes required for O-polysaccharide production. Conditional depletion of eipA expression in B. ovis results in a cell chaining phenotype, providing evidence that eipA directly or indirectly influences cell division in Brucella. We conclude that EipA is a molecular determinant of Brucella virulence that functions to maintain cell envelope integrity and influences cell division.

Brucella Periplasmic Protein EipB Is a Molecular Determinant of Cell Envelope Integrity and Virulence.

The Gram-negative cell envelope is a remarkable structure with core components that include an inner membrane, an outer membrane, and a peptidoglycan layer in the periplasmic space between. Multiple molecular systems function to maintain integrity of this essential barrier between the interior of the cell and its surrounding environment. We show that a conserved DUF1849 family protein, EipB, is secreted to the periplasmic space of Brucella species, a monophyletic group of intracellular pathogens. In the periplasm, EipB folds into an unusual 14-stranded ß-spiral structure that resembles the LolA and LolB lipoprotein delivery system, though the overall fold of EipB is distinct from LolA/LolB. Deletion of eipB results in defects in Brucella cell envelope integrity in vitro and in maintenance of spleen colonization in a mouse model of Brucella abortus infection. Transposon disruption of ttpA, which encodes a periplasmic protein containing tetratricopeptide repeats, is synthetically lethal with eipB deletion. ttpA is a reported virulence determinant in Brucella, and our studies of ttpA deletion and overexpression strains provide evidence that this gene also contributes to cell envelope function. We conclude that eipB and ttpA function in the Brucella periplasmic space to maintain cell envelope integrity, which facilitates survival in a mammalian host.IMPORTANCEBrucella species cause brucellosis, a global zoonosis. A gene encoding a conserved DUF1849-family protein, which we have named EipB, is present in all sequenced Brucella and several other genera in the class Alphaproteobacteria The manuscript provides the first functional and structural characterization of a DUF1849 protein. We show that EipB is secreted to the periplasm where it forms a spiral-shaped antiparallel ß protein that is a determinant of cell envelope integrity in vitro and virulence in an animal model of disease. eipB genetically interacts with ttpA, which also encodes a periplasmic protein. We propose that EipB and TtpA function as part of a system required for cell envelope homeostasis in select Alphaproteobacteria.

Molecular control of gene expression by Brucella BaaR, an IclR-type transcriptional repressor.

The general stress response sigma factor σE1 directly and indirectly regulates the transcription of dozens of genes that influence stress survival and host infection in the zoonotic pathogen Brucella abortus Characterizing the functions of σE1-regulated genes therefore would contribute to our understanding of B. abortus physiology and infection biology. σE1 indirectly activates transcription of the IclR family regulator Bab2_0215, but the function of this regulator remains undefined. Here, we present a structural and functional characterization of Bab2_0215, which we have named B rucella adipic acid-activated regulator (BaaR). We found that BaaR adopts a classic IclR-family fold and directly represses the transcription of two operons with predicted roles in carboxylic acid oxidation. BaaR binds two sites on chromosome II between baaR and a divergently transcribed hydratase/dehydrogenase (acaD2), and it represses transcription of both genes. We identified three carboxylic acids (adipic acid, tetradecanedioic acid, and ϵ-aminocaproic acid) and a lactone (ϵ-caprolactone) that enhance transcription from the baaR and acaD2 promoters. However, neither the activating acids nor caprolactone enhanced transcription by binding directly to BaaR. Induction of baaR transcription by adipic acid required the gene bab2_0213, which encodes a major facilitator superfamily transporter, suggesting that Bab2_0213 transports adipic acid across the inner membrane. We conclude that a suite of structurally related organic molecules activate transcription of genes repressed by BaaR. Our study provides molecular-level understanding of a gene expression program in B. abortus that is downstream of σE1.

Brucella abortus Induces a Warburg Shift in Host Metabolism That Is Linked to Enhanced Intracellular Survival of the Pathogen.

Intracellular bacterial pathogens exploit host cell resources to replicate and survive inside the host. Targeting these host systems is one promising approach to developing novel antimicrobials to treat intracellular infections. We show that human macrophage-like cells infected with Brucella abortus undergo a metabolic shift characterized by attenuated tricarboxylic acid cycle metabolism, reduced amino acid consumption, altered mitochondrial localization, and increased lactate production. This shift to an aerobic glycolytic state resembles the Warburg effect, a change in energy production that is well described in cancer cells and also occurs in activated inflammatory cells. B. abortus efficiently uses lactic acid as its sole carbon and energy source and requires the ability to metabolize lactate for normal survival in human macrophage-like cells. We demonstrate that chemical inhibitors of host glycolysis and lactate production do not affect in vitro growth of B. abortus in axenic culture but decrease its survival in the intracellular niche. Our data support a model in which infection shifts host metabolism to a Warburg-like state, and B. abortus uses this change in metabolism to promote intracellular survival. Pharmacological perturbation of these features of host cell metabolism may be a useful strategy to inhibit infection by intracellular pathogens.IMPORTANCEBrucella spp. are intracellular bacterial pathogens that cause disease in a range of mammals, including livestock. Transmission from livestock to humans is common and can lead to chronic human disease. Human macrophage-like cells infected with Brucella abortus undergo a Warburg-like metabolic shift to an aerobic glycolytic state where the host cells produce lactic acid and have reduced amino acid catabolism. We provide evidence that the pathogen can exploit this change in host metabolism to support growth and survival in the intracellular niche. Drugs that inhibit this shift in host cell metabolism inhibit intracellular replication and decrease the survival of B. abortus in an in vitro infection model; these drugs may be broadly useful therapeutics for intracellular infections.

Conserved ABC Transport System Regulated by the General Stress Response Pathways of Alpha- and Gammaproteobacteria.

Brucella abortus σE1 is an EcfG family sigma factor that regulates the transcription of dozens of genes in response to diverse stress conditions and is required for maintenance of chronic infection in a mouse model. A putative ATP-binding cassette transporter operon, bab1_0223-bab1_0226, is among the most highly activated gene sets in the σE1 regulon. The proteins encoded by the operon resemble quaternary ammonium-compatible solute importers but are most similar in sequence to the broadly conserved YehZYXW system, which remains largely uncharacterized. Transcription of yehZYXW is activated by the general stress sigma factor σS in Enterobacteriaceae, which suggests a functional role for this transport system in bacterial stress response across the classes Alphaproteobacteria and Gammaproteobacteria We present evidence that B. abortus YehZYXW does not function as an importer of known compatible solutes under physiological conditions and does not contribute to the virulence defect of a σE1-null strain. The sole in vitro phenotype associated with genetic disruption of this putative transport system is reduced growth in the presence of high Li+ ion concentrations. A crystal structure of B. abortus YehZ revealed a class II periplasmic binding protein fold with significant structural homology to Archaeoglobus fulgidus ProX, which binds glycine betaine. However, the structure of the YehZ ligand-binding pocket is incompatible with high-affinity binding to glycine betaine. This is consistent with weak measured binding of YehZ to glycine betaine and related compatible solutes. We conclude that YehZYXW is a conserved, stress-regulated transport system that is phylogenetically and functionally distinct from quaternary ammonium-compatible solute importers.IMPORTANCEBrucella abortus σE1 regulates transcription in response to stressors encountered in its mammalian host and is necessary for maintenance of chronic infection in a mouse model. The functions of the majority of genes regulated by σE1 remain undefined. We present a functional/structural analysis of a conserved putative membrane transport system (YehZYXW) whose expression is strongly activated by σE1 Though annotated as a quaternary ammonium osmolyte uptake system, experimental physiological studies and measured ligand-binding properties of the periplasmic binding protein (PBP), YehZ, are inconsistent with this function. A crystal structure of B. abortus YehZ provides molecular insight into differences between bona fide quaternary ammonium osmolyte importers and YehZ-related proteins, which form a distinct phylogenetic and functional group of PBPs.

WrpA Is an Atypical Flavodoxin Family Protein under Regulatory Control of the Brucella abortus General Stress Response System.

UNLABELLED: The general stress response (GSR) system of the intracellular pathogen Brucella abortus controls the transcription of approximately 100 genes in response to a range of stress cues. The core genetic regulatory components of the GSR are required for B. abortus survival under nonoptimal growth conditions in vitro and for maintenance of chronic infection in an in vivo mouse model. The functions of the majority of the genes in the GSR transcriptional regulon remain undefined. bab1_1070 is among the most highly regulated genes in this regulon: its transcription is activated 20- to 30-fold by the GSR system under oxidative conditions in vitro. We have solved crystal structures of Bab1_1070 and demonstrate that it forms a homotetrameric complex that resembles those of WrbA-type NADH:quinone oxidoreductases, which are members of the flavodoxin protein family. However, B. abortus WrbA-related protein (WrpA) does not bind flavin cofactors with a high affinity and does not function as an NADH:quinone oxidoreductase in vitro. Soaking crystals with flavin mononucleotide (FMN) revealed a likely low-affinity binding site adjacent to the canonical WrbA flavin binding site. Deletion of wrpA (ΔwrpA) does not compromise cell survival under acute oxidative stress in vitro or attenuate infection in cell-based or mouse models. However, a ΔwrpA strain does elicit increased splenomegaly in a mouse model, suggesting that WrpA modulates B. abortus interaction with its mammalian host. Despite high structural homology with canonical WrbA proteins, we propose that B. abortus WrpA represents a functionally distinct member of the diverse flavodoxin family. IMPORTANCE: Brucella abortus is an etiological agent of brucellosis, which is among the most common zoonotic diseases worldwide. The general stress response (GSR) regulatory system of B. abortus controls the transcription of approximately 100 genes and is required for maintenance of chronic infection in a murine model; the majority of GSR-regulated genes remain uncharacterized. We present in vitro and in vivo functional and structural analyses of WrpA, whose expression is strongly induced by GSR under oxidative conditions. Though WrpA is structurally related to NADH:quinone oxidoreductases, it does not bind redox cofactors in solution, nor does it exhibit oxidoreductase activity in vitro. However, WrpA does affect spleen inflammation in a murine infection model. Our data provide evidence that WrpA forms a new functional class of WrbA/flavodoxin family proteins.

Identification of a tissue-selective heat shock response regulatory network.

The heat shock response (HSR) is essential to survive acute proteotoxic stress and has been studied extensively in unicellular organisms and tissue culture cells, but to a lesser extent in intact metazoan animals. To identify the regulatory pathways that control the HSR in Caenorhabditis elegans, we performed a genome-wide RNAi screen and identified 59 genes corresponding to 7 positive activators required for the HSR and 52 negative regulators whose knockdown leads to constitutive activation of the HSR. These modifiers function in specific steps of gene expression, protein synthesis, protein folding, trafficking, and protein clearance, and comprise the metazoan heat shock regulatory network (HSN). Whereas the positive regulators function in all tissues of C. elegans, nearly all of the negative regulators exhibited tissue-selective effects. Knockdown of the subunits of the proteasome strongly induces HS reporter expression only in the intestine and spermatheca but not in muscle cells, while knockdown of subunits of the TRiC/CCT chaperonin induces HS reporter expression only in muscle cells. Yet, both the proteasome and TRiC/CCT chaperonin are ubiquitously expressed and are required for clearance and folding in all tissues. We propose that the HSN identifies a key subset of the proteostasis machinery that regulates the HSR according to the unique functional requirements of each tissue.

Phage against the Machine: The SIE-ence of Superinfection Exclusion.

Prophages can alter their bacterial hosts to prevent other phages from infecting the same cell, a mechanism known as superinfection exclusion (SIE). Such alterations are facilitated by phage interactions with critical bacterial components involved in motility, adhesion, biofilm production, conjugation, antimicrobial resistance, and immune evasion. Therefore, the impact of SIE extends beyond the immediate defense against superinfection, influencing the overall fitness and virulence of the bacteria. Evaluating the interactions between phages and their bacterial targets is critical for leading phage therapy candidates like Pseudomonas aeruginosa, a Gram-negative bacterium responsible for persistent and antibiotic-resistant opportunistic infections. However, comprehensive literature on the mechanisms underlying SIE remains scarce. Here, we provide a compilation of well-characterized and potential mechanisms employed by Pseudomonas phages to establish SIE. We hypothesize that the fitness costs imposed by SIE affect bacterial virulence, highlighting the potential role of this mechanism in the management of bacterial infections.

Identification of proteotoxic and proteoprotective bacteria that non-specifically affect proteins associated with neurodegenerative diseases.

There are no cures for neurodegenerative protein conformational diseases (PCDs), such as Alzheimer's disease (AD), Parkinson's disease (PD), and Huntington's disease (HD). Emerging evidence suggests the gut microbiota plays a role in their pathogenesis, though the influences of specific bacteria on disease-associated proteins remain elusive. Here, we reveal the effects of 229 human bacterial isolates on the aggregation and toxicity of Aß1-42, α-synuclein, and polyglutamine tracts in Caenorhabditis elegans expressing these culprit proteins. Our findings demonstrate that bacterial effects on host protein aggregation are consistent across different culprit proteins, suggesting that microbes affect protein stability by modulating host proteostasis rather than selectively targeting disease-associated proteins. Furthermore, we found that feeding C. elegans proteoprotective Prevotella corporis activates the heat shock response, revealing an unexpected discovery of a microbial influence on host proteostasis. Insight into how individual bacteria affect PCD proteins could open new strategies for prevention and treatment by altering the abundance of microbes.

Whole-genome sequencing and analysis of Streptococcus iniae strain isolated from the brain of tilapia (Oreochromis sp.).

Streptococcus iniae is a bacterium that can infect fish, mammals, and humans. In this study, the S. iniae-313 strain was isolated from the brain of an infected tilapia, and the analysis of its sequenced genome is reported. The data revealed that S. iniae-313 carried antibiotic-resistant genes and virulence factors.

Quantification of Bacterial Loads in Caenorhabditis elegans.

Caenorhabditis elegans is a ubiquitous free-living nematode that feeds on bacteria. The organism was introduced into a laboratory setting in the 1970s and has since gained popularity as a model to study host-bacteria interactions. One advantage of using C. elegans is that its intestine can be colonized by the bacteria on which it feeds. Quantifying the bacterial load within C. elegans is an important and easily obtainable metric when investigating host-bacteria interactions. Although quantification of bacteria harbored in C. elegans via whole-worm lysis is not a novel assay, there is great variation between existing methods. To lyse C. elegans, many protocols rely on the use of a hand-held homogenizer, which could introduce systematic error and subsequent variation between researchers performing the same experiment. Here, we describe a method of lysing the intestines of C. elegans to quantify the bacterial load within the intestine. Our method has been optimized for removing exogenous bacteria while maintaining worm paralysis, to ensure no bactericidal agents are swallowed, which could kill bacteria within the intestine and affect results. We utilize and compare the efficiency of two different homogenization tools: a battery-powered hand-held homogenizer, and a benchtop electric homogenizer, where the latter minimizes variability. Thus, our protocol has been optimized to reduce systematic error and decrease the potential for variability among experimenters. Graphic abstract: Simplified overview of the procedure used to quantify the bacterial load within C. elegans. The two different methods are herein described for worm lysis: "Option 1" is a hand-held homogenizer, and "Option 2" is a benchtop homogenizer.

Time-off-pick Assay to Measure Caenorhabditis elegans Motility.

Caenorhabditis elegans is a simple metazoan that is often used as a model organism to study various human ailments with impaired motility phenotypes, including protein conformational diseases. Numerous motility assays that measure neuro-muscular function have been employed using C. elegans . Here, we describe "time-off-pick" (TOP), a novel assay for assessing motility in C. elegans . TOP is conducted by sliding an eyebrow hair under the mid-section of the worm and counting the number of seconds it takes for the worm to crawl completely off. The time it takes for the worm to crawl off the eyebrow hair is proportional to the severity of its motility defect. Other readouts of motility include crawling or swimming phenotypes, and although widely established, have some limitations. For example, worms that are roller mutants are less suitable for crawling or swimming assays. We demonstrated that our novel TOP assay is sensitive to age-dependent changes in motility, thus, providing another more inclusive method to assess motor function in C. elegans . Graphical abstract: Conceptual overview of the "time-off-pick" (TOP) assay. Various C. elegans models exhibit age-dependent defects in motility. The time it takes for a worm to crawl off of an eyebrow pick that is slid under its mid-section is measured in TOP seconds. A greater TOP is indicative of a greater motility defect. Eventually, worms with phenotypes that lead to paralysis will not be able to leave the pick.

Silver nanoparticles enhance the efficacy of aminoglycosides against antibiotic-resistant bacteria.

As the threat of antimicrobial-resistant bacteria compromises the safety and efficacy of modern healthcare practices, the search for effective treatments is more urgent than ever. For centuries, silver (Ag) has been known to have antibacterial properties and, over the past two decades, Ag-based nanoparticles have gained traction as potential antimicrobials. The antibacterial efficacy of Ag varies with structure, size, and concentration. In the present study, we examined Ag nanoparticles (AgNPs) for their antimicrobial activity and safety. We compared different commercially-available AgNPs against gram-negative Escherichia coli, Pseudomonas aeruginosa, Acinetobacter baumannii, and gram-positive Staphylococcus aureus methicillin-resistant and susceptible strains. The most effective formula of AgNPs tested had single-digit (µg/mL) minimum inhibitory concentrations against gram-negative multidrug-resistant clinical bacterial isolates with novel and emerging mechanisms of resistance. The mode of killing was assessed in E. coli and was found to be bactericidal, which is consistent with previous studies using other AgNP formulations. We evaluated cytotoxicity by measuring physiological readouts using the Caenorhabditis elegans model and found that motility was affected, but not the lifespan. Furthermore, we found that at their antibacterial concentrations, AgNPs were non-cytotoxic to any of the mammalian cell lines tested, including macrophages, stem cells, and epithelial cells. More interestingly, our experiments revealed synergy with clinically relevant antibiotics. We found that a non-toxic and non-effective concentration of AgNPs reduced the minimum inhibitory concentrations of aminoglycoside by approximately 22-fold. Because both aminoglycosides and Ag are known to target the bacterial ribosome, we tested whether Ag could also target eukaryotic ribosomes. We measured the rate of mistranslation at bactericidal concentration and found no effect, indicating that AgNPs are not proteotoxic to the host at the tested concentrations. Collectively, our results suggest that AgNPs could have a promising clinical application as a potential stand-alone therapy or antibiotic adjuvants.

The potential impact of the COVID-19 pandemic on antimicrobial resistance and antibiotic stewardship.

The rapid emergence and spread of antimicrobial resistance continue to kill an estimated 700,000 people annually, and this number is projected to increase ten-fold by 2050. With the lack of data, it is uncertain how the COVID-19 pandemic will affect antimicrobial resistance. Severe disruption of research, innovation, global health programs, and compromised antimicrobial stewardship, infection prevention and control programs, especially in low-and middle-income countries, could affect antimicrobial resistance. However, factors such as strict lockdown, social distancing, vaccination, and the extensive implementation of hand hygiene and face masks, with limited international travel and migration, may also contribute to decreasing AMR. Although the impact of COVID-19 on AMR is global, the adverse effect is likely to be worse in LMICs. In this article, we explore the possible impact of the current pandemic on antibiotic resistance.

The gigA/gigB Genes Regulate the Growth, Stress Response, and Virulence of Acinetobacter baumannii ATCC 17978 Strain.

Acinetobacter baumannii is an important pathogen of nosocomial infection. Recently, a group of genes, named "gig" (for Growth in Galleria), have been identified in a contemporary multi-drug resistant clinical isolate of A. baumannii-strain AB5075. Among these so-called gig genes, gigA and gigB were found to promote antibiotic resistance, stress survival, and virulence of AB5075 by interacting with the nitrogen phosphotransferase system (PTSNtr). This study aimed to investigate the roles of gigA/gigB, which appear to comprise a stress-signaling pathway (encoding for an atypical two-component system response regulator and a predicted anti-anti-sigma factor, respectively), and the involvement of ptsP (encoding the Enzyme I component of the PTSNtr) in the growth, stress resistance, and virulence of the widely studied A. baumannii strain ATCC 17978. Genetic analyses of strains harboring mutations of gigA and gigB were performed to investigate the roles of these genes in bacterial growth, stress resistance, evading macrophage defense, and killing of Galleria mellonella larva. In contrast with findings from strain AB5075 where gigA and gigB contribute to aminoglycoside resistance, the data presented herein indicate that the loss of gigA/gigB does not impact antibiotic resistance of strain ATCC 17978. Interestingly, however, we found that deletion of gigA/gigB in the ATCC 17978 background imparts a general growth in laboratory medium and also conferred growth and replication defects within murine macrophages and an inability to kill G. mellonella larvae. Importantly, studies as well as the loss of ptsP restored the phenotypes of the gigA/gigB mutant to that of the wild-type. The data presented herein indicate that in A. baumannii ATCC 17978, the gigA/gigB genes play a key role in both growth and virulence traits, but are dispensable for other stress-resistance survival phenotypes, including aminoglycoside resistance. Our findings thus highlight several similarities and also important differences between the gigA/gigB stress-signaling pathway in two commonly studied isolates of this troublesome pathogen.