Vaccines against extraintestinal pathogenic Escherichia coli (ExPEC): progress and challenges.

The emergence of antimicrobial resistance (AMR) is a principal global health crisis projected to cause 10 million deaths annually worldwide by 2050. While the Gram-negative bacteria Escherichia coli is commonly found as a commensal microbe in the human gut, some strains are dangerously pathogenic, contributing to the highest AMR-associated mortality. Strains of E. coli that can translocate from the gastrointestinal tract to distal sites, called extraintestinal E. coli (ExPEC), are particularly problematic and predominantly afflict women, the elderly, and immunocompromised populations. Despite nearly 40 years of clinical trials, there is still no vaccine against ExPEC. One reason for this is the remarkable diversity in the ExPEC pangenome across pathotypes, clades, and strains, with hundreds of genes associated with pathogenesis including toxins, adhesins, and nutrient acquisition systems. Further, ExPEC is intimately associated with human mucosal surfaces and has evolved creative strategies to avoid the immune system. This review summarizes previous and ongoing preclinical and clinical ExPEC vaccine research efforts to help identify key gaps in knowledge and remaining challenges.

Progress toward a vaccine for extraintestinal pathogenic E. coli (ExPEC) II: efficacy of a toxin-autotransporter dual antigen approach.

Extraintestinal pathogenic Escherichia coli (ExPEC) is a leading cause of worldwide morbidity and mortality, the top cause of antimicrobial-resistant (AMR) infections, and the most frequent cause of life-threatening sepsis and urinary tract infections (UTI) in adults. The development of an effective and universal vaccine is complicated by this pathogen's pan-genome, its ability to mix and match virulence factors and AMR genes via horizontal gene transfer, an inability to decipher commensal from pathogens, and its intimate association and co-evolution with mammals. Using a pan virulome analysis of >20,000 sequenced E. coli strains, we identified the secreted cytolysin α-hemolysin (HlyA) as a high priority target for vaccine exploration studies. We demonstrate that a catalytically inactive pure form of HlyA, expressed in an autologous host using its own secretion system, is highly immunogenic in a murine host, protects against several forms of ExPEC infection (including lethal bacteremia), and significantly lowers bacterial burdens in multiple organ systems. Interestingly, the combination of a previously reported autotransporter (SinH) with HlyA was notably effective, inducing near complete protection against lethal challenge, including commonly used infection strains ST73 (CFT073) and ST95 (UTI89), as well as a mixture of 10 of the most highly virulent sequence types and strains from our clinical collection. Both HlyA and HlyA-SinH combinations also afforded some protection against UTI89 colonization in a murine UTI model. These findings suggest recombinant, inactive hemolysin and/or its combination with SinH warrant investigation in the development of an E. coli vaccine against invasive disease.

Assessment of a SARS-CoV-2 wastewater monitoring program in El Paso, Texas, from November 2020 to June 2022.

The border city of El Paso, Texas, and its water utility, El Paso Water, initiated a SARS-CoV-2 wastewater monitoring program to assess virus trends and the appropriateness of a wastewater monitoring program for the community. Nearly weekly sample collection at four wastewater treatment facilities (WWTFs), serving distinct regions of the city, was analyzed for SARS-CoV-2 genes using the CDC 2019-Novel coronavirus Real-Time RT-PCR diagnostic panel. Virus concentrations ranged from 86.7 to 268,000 gc/L, varying across time and at each WWTF. The lag time between virus concentrations in wastewater and reported COVID-19 case rates (per 100,00 population) ranged from 4-24 days for the four WWTFs, with the strongest trend occurring from November 2021 - June 2022. This study is an assessment of the utility of a geographically refined SARS-CoV-2 wastewater monitoring program to supplement public health efforts that will manage the virus as it becomes endemic in El Paso.

Wastewater sequencing reveals community and variant dynamics of the collective human virome.

Wastewater is a discarded human by-product, but its analysis may help us understand the health of populations. Epidemiologists first analyzed wastewater to track outbreaks of poliovirus decades ago, but so-called wastewater-based epidemiology was reinvigorated to monitor SARS-CoV-2 levels while bypassing the difficulties and pit falls of individual testing. Current approaches overlook the activity of most human viruses and preclude a deeper understanding of human virome community dynamics. Here, we conduct a comprehensive sequencing-based analysis of 363 longitudinal wastewater samples from ten distinct sites in two major cities. Critical to detection is the use of a viral probe capture set targeting thousands of viral species or variants. Over 450 distinct pathogenic viruses from 28 viral families are observed, most of which have never been detected in such samples. Sequencing reads of established pathogens and emerging viruses correlate to clinical data sets of SARS-CoV-2, influenza virus, and monkeypox viruses, outlining the public health utility of this approach. Viral communities are tightly organized by space and time. Finally, the most abundant human viruses yield sequence variant information consistent with regional spread and evolution. We reveal the viral landscape of human wastewater and its potential to improve our understanding of outbreaks, transmission, and its effects on overall population health.

Class-Driven Synergy and Antagonism between a Pseudomonas Phage and Antibiotics.

The ubiquitous bacterial pathogen Pseudomonas aeruginosa is responsible for severe infections in patients with burns, cystic fibrosis, and neutropenia. Biofilm formation gives physical refuge and a protected microenvironment for sessile cells, rendering cure by antibiotics a challenge. Bacteriophages have evolved to prey on these biofilms over millions of years, using hydrolases and depolymerases to penetrate biofilms and reach cellular targets. Here, we assessed how a newly discovered KMV-like phage (ΦJB10) interacts with antibiotics to treat P. aeruginosa more effectively in both planktonic and biofilm forms. By testing representatives of four classes of antibiotics (cephalosporins, aminoglycosides, fluoroquinolones, and carbapenems), we demonstrated class-dependent interactions between ΦJB10 and antibiotics in both biofilm clearance and P. aeruginosa killing. Despite identifying antagonism between some antibiotic classes and ΦJB10 at early time points, all classes showed neutral to favorable interactions with the phage at later time points. In one notable example where the antibiotic alone had poor activity against both biofilm and high-density planktonic cells, we found that addition of ΦJB10 demonstrated synergy and resulted in effective treatment of both. Further, ΦJB10 seemed to act as an adjuvant to several antibiotics, reducing the concentration of antibiotics required to ablate the biofilm. This report shows that phages such as ΦJB10 may be valuable additions to the armamentarium against difficult-to-treat biofilm-based infections.

High levels of lipoprotein(a) in transgenic mice exacerbate atherosclerosis and promote vulnerable plaque features in a sex-specific manner.

BACKGROUND AND AIMS: Despite increased clinical interest in lipoprotein(a) (Lp(a)), many questions remain about the molecular mechanisms by which it contributes to atherosclerotic cardiovascular disease. Existing murine transgenic (Tg) Lp(a) models are limited by low plasma levels of Lp(a) and have not consistently shown a pro-atherosclerotic effect of Lp(a). METHODS: We generated Tg mice expressing both human apolipoprotein(a) (apo(a)) and human apoB-100, with pathogenic levels of plasma Lp(a) (range 87-250 mg/dL). Female and male Lp(a) Tg mice (Tg(LPA+/0;APOB+/0)) and human apoB-100-only controls (Tg(APOB+/0)) (n = 10-13/group) were fed a high-fat, high-cholesterol diet for 12 weeks, with Ldlr knocked down using an antisense oligonucleotide. FPLC was used to characterize plasma lipoprotein profiles. Plaque area and necrotic core size were quantified and immunohistochemical assessment of lesions using a variety of cellular and protein markers was performed. RESULTS: Male and female Tg(LPA+/0;APOB+/0) and Tg(APOB+/0) mice exhibited proatherogenic lipoprotein profiles with increased cholesterol-rich VLDL and LDL-sized particles and no difference in plasma total cholesterol between genotypes. Complex lesions developed in the aortic sinus of all mice. Plaque area (+22%), necrotic core size (+25%), and calcified area (+65%) were all significantly increased in female Tg(LPA+/0;APOB+/0) mice compared to female Tg(APOB+/0) mice. Immunohistochemistry of lesions demonstrated that apo(a) deposited in a similar pattern as apoB-100 in Tg(LPA+/0;APOB+/0) mice. Furthermore, female Tg(LPA+/0;APOB+/0) mice exhibited less organized collagen deposition as well as 42% higher staining for oxidized phospholipids (OxPL) compared to female Tg(APOB+/0) mice. Tg(LPA+/0;APOB+/0) mice had dramatically higher levels of plasma OxPL-apo(a) and OxPL-apoB compared to Tg(APOB+/0) mice, and female Tg(LPA+/0;APOB+/0) mice had higher plasma levels of the proinflammatory cytokine MCP-1 (+3.1-fold) compared to female Tg(APOB+/0) mice. CONCLUSIONS: These data suggest a pro-inflammatory phenotype exhibited by female Tg mice expressing Lp(a) that appears to contribute to the development of more severe lesions with greater vulnerable features.

A Retrospective, Observational Study of 12 Cases of Expanded-Access Customized Phage Therapy: Production, Characteristics, and Clinical Outcomes.

BACKGROUND: Antimicrobial resistance (AMR) is undermining modern medicine, a problem compounded by bacterial adaptation to antibiotic pressures. Phages are viruses that infect bacteria. Their diversity and evolvability offer the prospect of their use as a therapeutic solution. Reported are outcomes of customized phage therapy for patients with difficult-to-treat antimicrobial resistant infections. METHODS: We retrospectively assessed 12 cases of customized phage therapy from a phage production center. Phages were screened, purified, sequenced, characterized, and Food and Drug Administration-approved via the IND (investigational new drug) compassionate-care route. Outcomes were assessed as favorable or unfavorable by microbiologic and clinical standards. Infections were device-related or systemic. Other experiences such as time to treatment, antibiotic synergy, and immune responses were recorded. RESULTS: Fifty requests for phage therapy were received. Customized phages were generated for 12 patients. After treatment, 42% (5/12) of cases showed bacterial eradication and 58% (7/12) showed clinical improvement, with two-thirds of all cases (66%) showing favorable responses. No major adverse reactions were observed. Antibiotic-phage synergy in vitro was observed in most cases. Immunological neutralization of phages was reported in 5 cases. Several cases were complicated by secondary infections. Complete characterization of the phages (morphology, genomics, and activity) and their production (methods, sterility, and endotoxin tests) are reported. CONCLUSIONS: Customized phage production and therapy was safe and yielded favorable clinical or microbiological outcomes in two-thirds of cases. A center or pipeline dedicated to tailoring the phages against a patient's specific AMR bacterial infection may be a viable option where standard treatment has failed.

Wastewater pandemic preparedness: Toward an end-to-end pathogen monitoring program.

Molecular analysis of public wastewater has great potential as a harbinger for community health and health threats. Long-used to monitor the presence of enteric viruses, in particular polio, recent successes of wastewater as a reliable lead indicator for trends in SARS-CoV-2 levels and hospital admissions has generated optimism and emerging evidence that similar science can be applied to other pathogens of pandemic potential (PPPs), especially respiratory viruses and their variants of concern (VOC). However, there are substantial challenges associated with implementation of this ideal, namely that multiple and distinct fields of inquiry must be bridged and coordinated. These include engineering, molecular sciences, temporal-geospatial analytics, epidemiology and medical, and governmental and public health messaging, all of which present their own caveats. Here, we outline a framework for an integrated, state-wide, end-to-end human pathogen monitoring program using wastewater to track viral PPPs.

Broad protective vaccination against systemic Escherichia coli with autotransporter antigens.

Extraintestinal pathogenic Escherichia coli (ExPEC) is the leading cause of adult life-threatening sepsis and urinary tract infections (UTI). The emergence and spread of multidrug-resistant (MDR) ExPEC strains result in a considerable amount of treatment failure and hospitalization costs, and contribute to the spread of drug resistance amongst the human microbiome. Thus, an effective vaccine against ExPEC would reduce morbidity and mortality and possibly decrease carriage in healthy or diseased populations. A comparative genomic analysis demonstrated a gene encoding an invasin-like protein, termed sinH, annotated as an autotransporter protein, shows high prevalence in various invasive ExPEC phylogroups, especially those associated with systemic bacteremia and UTI. Here, we evaluated the protective efficacy and immunogenicity of a recombinant SinH-based vaccine consisting of either domain-3 or domains-1,2, and 3 of the putative extracellular region of surface-localized SinH. Immunization of a murine host with SinH-based antigens elicited significant protection against various strains of the pandemic ExPEC sequence type 131 (ST131) as well as multiple sequence types in two distinct models of infection (colonization and bacteremia). SinH immunization also provided significant protection against ExPEC colonization in the bladder in an acute UTI model. Immunized cohorts produced significantly higher levels of vaccine-specific serum IgG and urinary IgG and IgA, findings consistent with mucosal protection. Collectively, these results demonstrate that autotransporter antigens such as SinH may constitute promising ExPEC phylogroup-specific and sequence-type effective vaccine targets that reduce E. coli colonization and virulence.

Genomic and Functional Characterization of Vancomycin-Resistant Enterococci-Specific Bacteriophages in the Galleria mellonella Wax Moth Larvae Model.

Phages are naturally occurring viruses that selectively kill bacterial species without disturbing the individual's normal flora, averting the collateral damage of antimicrobial usage. The safety and the effectiveness of phages have been mainly confirmed in the food industry as well as in animal models. In this study, we report on the successful isolation of phages specific to Vancomycin-resistant Enterococci, including Enterococcus faecium (VREfm) and Enterococcus faecalis from sewage samples, and demonstrate their efficacy and safety for VREfm infection in the greater wax moth Galleria mellonella model. No virulence-associated genes, antibiotic resistance genes or integrases were detected in the phages' genomes, rendering them safe to be used in an in vivo model. Phages may be considered as potential agents for therapy for bacterial infections secondary to multidrug-resistant organisms such as VREfm.

Phage Resistance Accompanies Reduced Fitness of Uropathogenic Escherichia coli in the Urinary Environment.

Urinary tract infection (UTI) is among the most common infections treated worldwide each year and is caused primarily by uropathogenic Escherichia coli (UPEC). Rising rates of antibiotic resistance among uropathogens have spurred a consideration of alternative treatment strategies, such as bacteriophage (phage) therapy; however, phage-bacterial interactions within the urinary environment are poorly defined. Here, we assess the activity of two phages, namely, HP3 and ES17, against clinical UPEC isolates using in vitro and in vivo models of UTI. In both bacteriologic medium and pooled human urine, we identified phage resistance arising within the first 6 to 8 h of coincubation. Whole-genome sequencing revealed that UPEC strains resistant to HP3 and ES17 harbored mutations in genes involved in lipopolysaccharide (LPS) biosynthesis. Phage-resistant strains displayed several in vitro phenotypes, including alterations to adherence to and invasion of human bladder epithelial HTB-9 cells and increased biofilm formation in some isolates. Interestingly, these phage-resistant UPEC isolates demonstrated reduced growth in pooled human urine, which could be partially rescued by nutrient supplementation and were more sensitive to several outer membrane-targeting antibiotics than parental strains. Additionally, phage-resistant UPEC isolates were attenuated in bladder colonization in a murine UTI model. In total, our findings suggest that while resistance to phages, such as HP3 and ES17, may arise readily in the urinary environment, phage resistance is accompanied by fitness costs which may render UPEC more susceptible to host immunity or antibiotics. IMPORTANCE UTI is one of the most common causes of outpatient antibiotic use, and rising antibiotic resistance threatens the ability to control UTI unless alternative treatments are developed. Bacteriophage (phage) therapy is gaining renewed interest; however, much like with antibiotics, bacteria can readily become resistant to phages. For successful UTI treatment, we must predict how bacteria will evade killing by phage and identify the downstream consequences of phage resistance during bacterial infection. In our current study, we found that while phage-resistant bacteria quickly emerged in vitro, these bacteria were less capable of growing in human urine and colonizing the murine bladder. These results suggest that phage therapy poses a viable UTI treatment if phage resistance confers fitness costs for the uropathogen. These results have implications for developing cocktails of phage with multiple different bacterial targets, of which each is evaded only at the cost of bacterial fitness.

Corrigendum: Development of phage cocktails to treat E. coli catheter-associated urinary tract infection and associated biofilms.

[This corrects the article DOI: 10.3389/fmicb.2022.796132.].

Development of Phage Cocktails to Treat E. coli Catheter-Associated Urinary Tract Infection and Associated Biofilms.

High rates of antimicrobial resistance and formation of biofilms makes treatment of Escherichia coli catheter-associated urinary tract infections (CAUTI) particularly challenging. CAUTI affect 1 million patients per year in the United States and are associated with morbidity and mortality, particularly as an etiology for sepsis. Phage have been proposed as a potential therapeutic option. Here, we report the development of phage cocktails that lyse contemporary E. coli strains isolated from the urine of patients with spinal cord injury (SCI) and display strong biofilm-forming properties. We characterized E. coli phage against biofilms in two in vitro CAUTI models. Biofilm viability was measured by an MTT assay that determines cell metabolic activity and by quantification of colony forming units. Nine phage decreased cell viability by >80% when added individually to biofilms of two E. coli strains in human urine. A phage cocktail comprising six phage lyses 82% of the strains in our E. coli library and is highly effective against young and old biofilms and against biofilms on silicon catheter materials. Using antibiotics together with our phage cocktail prevented or decreased emergence of E. coli resistant to phage in human urine. We created an anti-biofilm phage cocktail with broad host range against E. coli strains isolated from urine. These phage cocktails may have therapeutic potential against CAUTI.

Sortilin enhances secretion of apolipoprotein(a) through effects on apolipoprotein B secretion and promotes uptake of lipoprotein(a).

Elevated plasma lipoprotein(a) (Lp(a)) is an independent, causal risk factor for atherosclerotic cardiovascular disease and calcific aortic valve stenosis. Lp(a) is formed in or on hepatocytes from successive noncovalent and covalent interactions between apo(a) and apoB, although the subcellular location of these interactions and the nature of the apoB-containing particle involved remain unclear. Sortilin, encoded by the SORT1 gene, modulates apoB secretion and LDL clearance. We used a HepG2 cell model to study the secretion kinetics of apo(a) and apoB. Overexpression of sortilin increased apo(a) secretion, while siRNA-mediated knockdown of sortilin expression correspondingly decreased apo(a) secretion. Sortilin binds LDL but not apo(a) or Lp(a), indicating that its effect on apo(a) secretion is likely indirect. Indeed, the effect was dependent on the ability of apo(a) to interact noncovalently with apoB. Overexpression of sortilin enhanced internalization of Lp(a), but not apo(a), by HepG2 cells, although neither sortilin knockdown in these cells or Sort1 deficiency in mice impacted Lp(a) uptake. We found several missense mutations in SORT1 in patients with extremely high Lp(a) levels; sortilin containing some of these mutations was more effective at promoting apo(a) secretion than WT sortilin, though no differences were found with respect to Lp(a) internalization. Our observations suggest that sortilin could play a role in determining plasma Lp(a) levels and corroborate in vivo human kinetic studies which imply that secretion of apo(a) and apoB are coupled, likely within the hepatocyte.

Apo(a) and ApoB Interact Noncovalently Within Hepatocytes: Implications for Regulation of Lp(a) Levels by Modulation of ApoB Secretion.

BACKGROUND: Elevated plasma Lp(a) (lipoprotein(a)) levels are associated with increased risk for atherosclerotic cardiovascular disease and aortic valve stenosis. However, the cell biology of Lp(a) biosynthesis remains poorly understood, with the locations of the noncovalent and covalent steps of Lp(a) assembly unclear and the nature of the apoB-containing particle destined for Lp(a) unknown. We, therefore, asked if apo(a) and apoB interact noncovalently within hepatocytes and if this impacts Lp(a) biosynthesis. METHODS: Using human hepatocellular carcinoma cells expressing 17K (17 kringle) apo(a), or a 17KΔLBS7,8 variant with a reduced ability to bind noncovalently to apoB, we performed coimmunoprecipitation, coimmunofluorescence, and proximity ligation assays to document intracellular apo(a):apoB interactions. We used a pulse-chase metabolic labeling approach to measure apo(a) and apoB secretion rates. RESULTS: Noncovalent complexes containing apo(a)/apoB are present in lysates from cells expressing 17K but not 17KΔLBS7,8, whereas covalent apo(a)/apoB complexes are absent from lysates. 17K and apoB colocalized intracellularly, overlapping with staining for markers of endoplasmic reticulum trans-Golgi, and early endosomes, and less so with lysosomes. The 17KΔLBS7,8 had lower colocalization with apoB. Proximity ligation assays directly documented intracellular 17K/apoB interactions, which were dramatically reduced for 17KΔLBS7,8. Treatment of cells with PCSK9 (proprotein convertase subtilisin/kexin type 9) enhanced, and lomitapide reduced, apo(a) secretion in a manner dependent on the noncovalent interaction between apo(a) and apoB. Apo(a) secretion was also reduced by siRNA-mediated knockdown of APOB. CONCLUSIONS: Our findings explain the coupling of apo(a) and Lp(a)-apoB production observed in human metabolic studies using stable isotopes as well as the ability of agents that inhibit apoB biosynthesis to lower Lp(a) levels.

Development of Antimicrobial Nitric Oxide-Releasing Fibers.

Nitric oxide (NO) is a highly reactive gas molecule, exhibiting antimicrobial properties. Because of its reactive nature, it is challenging to store and deliver NO efficiently as a therapeutic agent. The objective of this study was to develop NO-releasing polymeric fibers (NO-fibers), as an effective delivery platform for NO. NO-fibers were fabricated with biopolymer solutions of polyvinyl pyrrolidone (PVP) and ethylcellulose (EC), and derivatives of N-diazeniumdiolate (NONOate) as NO donor molecules, using an electrospinning system. We evaluated in vitro NO release kinetics, along with antimicrobial effects and cytotoxicity in microorganisms and human cell culture models. We also studied the long-term stability of NONOates in NO-fibers over 12 months. We demonstrated that the NO-fibers could release NO over 24 h, and showed inhibition of the growth of Pseudomonas aeruginosa (P. aeruginosa) and methicillin-resistant Staphylococcus aureus (MRSA), without causing cytotoxicity in human cells. NO-fibers were able to store NONOates for over 12 months at room temperature. This study presents the development of NO-fibers, and the feasibility of NO-fibers to efficiently store and deliver NO, which can be further developed as a bandage.

Antiviral Resistance and Phage Counter Adaptation to Antibiotic-Resistant Extraintestinal Pathogenic Escherichia coli.

Extraintestinal pathogenic Escherichia coli (ExPEC), often multidrug resistant (MDR), is a leading cause of urinary tract and systemic infections. The crisis of emergent MDR pathogens has led some to propose bacteriophages as a therapeutic. However, bacterial resistance to phage is a concerning issue that threatens to undermine phage therapy. Here, we demonstrate that E. coli sequence type 131, a circulating pandemic strain of ExPEC, rapidly develops resistance to a well-studied and therapeutically active phage (ϕHP3). Whole-genome sequencing of the resisters revealed truncations in genes involved in lipopolysaccharide (LPS) biosynthesis, the outer membrane transporter ompA, or both, implicating them as phage receptors. We found ExPEC resistance to phage is associated with a loss of fitness in host microenvironments and attenuation in a murine model of systemic infection. Furthermore, we constructed a novel phage-bacterium bioreactor to generate an evolved phage isolate with restored infectivity to all LPS-truncated ExPEC resisters. This study suggests that although the resistance of pandemic E. coli to phage is frequent, it is associated with attenuation of virulence and susceptibility to new phage variants that arise by directed evolution.IMPORTANCE In response to the rising crisis of antimicrobial resistance, bacteriophage (phage) therapy has gained traction. In the United States, there have been over 10 cases of largely successful compassionate-use phage therapy to date. The resilience of pathogens allowing their broad antibiotic resistance means we must also consider resistance to therapeutic phages. This work fills gaps in knowledge regarding development of phage resisters in a model of infection and finds critical fitness losses in those resisters. We also found that the phage was able to rapidly readapt to these resisters.

Targeting of Mammalian Glycans Enhances Phage Predation in the Gastrointestinal Tract.

The human gastrointestinal mucosal surface consists of a eukaryotic epithelium, a prokaryotic microbiota, and a carbohydrate-rich interface that separates them. In the gastrointestinal tract, the interaction of bacteriophages (phages) and their prokaryotic hosts influences the health of the mammalian host, especially colonization with invasive pathobionts. Antibiotics may be used, but they also kill protective commensals. Here, we report a novel phage whose lytic cycle is enhanced in intestinal environments. The tail fiber gene, whose protein product binds human heparan sulfated proteoglycans and localizes the phage to the epithelial cell surface, positions it near its bacterial host, a type of locational targeting mechanism. This finding offers the prospect of developing mucosal targeting phage to selectively remove invasive pathobiont species from mucosal surfaces.IMPORTANCE Invasive pathobionts or microbes capable of causing disease can reside deep within the mucosal epithelium of our gastrointestinal tract. Targeted effective antibacterial therapies are needed to combat these disease-causing organisms, many of which may be multidrug resistant. Here, we isolated a lytic bacteriophage (phage) that can localize to the epithelial surface by binding heparan sulfated glycans, positioning it near its host, Escherichia coli This targeted therapy can be used to selectively remove invasive pathobionts from the gastrointestinal tract, preventing the development of disease.

Lipoprotein(a): Expanding our knowledge of aortic valve narrowing.

Elevated levels of lipoprotein(a) [Lp(a)] have been identified as an independent and causal risk factor for atherosclerotic cardiovascular disease (ASCVD) and, more recently, calcific aortic valve disease (CAVD). CAVD is a slow, progressive disorder presenting as severe trileaflet calcification known as aortic valve stenosis (AS) that impairs valve motion and restricts ventricular outflow. AS afflicts 2% of the aging population (≥ 65 years) and tends to be quite advanced by the time it presents clinical symptoms of exertional angina, syncope, or heart failure. Currently, the only effective clinical therapy for AS patients is surgical or transcatheter aortic valve replacement. Evidence is accumulating that Lp(a) can exacerbate pathophysiological processes in CAVD, specifically, endothelial dysfunction, formation of foam cells, and promotion of a pro-inflammatory state. In the valve milieu, the pro-inflammatory effects of Lp(a) are manifested in valve thickening and mineralization through pro-osteogenic signaling and changes in gene expression in valve interstitial cells that is primarily facilitated by the oxidized phospholipid content of Lp(a). In AS pathogenesis, an incomplete understanding of the role of Lp(a) at the molecular level and the absence of appropriate animal models are barriers for the development of specific and effective clinical interventions designed to mitigate the role of Lp(a) in AS. However, the advent of effective therapies that dramatically lower Lp(a) provides the possibility of the first medical treatment to halt AS progression.

Enteroaggregative E. coli Adherence to Human Heparan Sulfate Proteoglycans Drives Segment and Host Specific Responses to Infection.

Enteroaggregative Escherichia coli (EAEC) is a significant cause of acute and chronic diarrhea, foodborne outbreaks, infections of the immunocompromised, and growth stunting in children in developing nations. There is no vaccine and resistance to antibiotics is rising. Unlike related E. coli pathotypes that are often associated with acute bouts of infection, EAEC is associated with persistent diarrhea and subclinical long-term colonization. Several secreted virulence factors have been associated with EAEC pathogenesis and linked to disease in humans, less certain are the molecular drivers of adherence to the intestinal mucosa. We previously established human intestinal enteroids (HIEs) as a model system to study host-EAEC interactions and aggregative adherence fimbriae A (AafA) as a major driver of EAEC adherence to HIEs. Here, we report a large-scale assessment of the host response to EAEC adherence from all four segments of the intestine across at least three donor lines for five E. coli pathotypes. The data demonstrate that the host response in the duodenum is driven largely by the infecting pathotype, whereas the response in the colon diverges in a patient-specific manner. Major pathways altered in gene expression in each of the four enteroid segments differed dramatically, with responses observed for inflammation, apoptosis and an overwhelming response to different mucin genes. In particular, EAEC both associated with large mucus droplets and specific mucins at the epithelial surface, binding that was ameliorated when mucins were removed, a process dependent on AafA. Pan-screening for glycans for binding to purified AafA identified the human ligand as heparan sulfate proteoglycans (HSPGs). Removal of HSPG abrogated EAEC association with HIEs. These results may mean that the human intestine responds remarkably different to distinct pathobionts that is dependent on the both the individual and intestinal segment in question, and uncover a major role for surface heparan sulfate proteoglycans as tropism-driving factor in adherence and/or colonization.