Distinct Escherichia coli transcriptional profiles in the guts of recurrent UTI sufferers revealed by pangenome hybrid selection.

Low-abundance members of microbial communities are difficult to study in their native habitats. This includes Escherichia coli, a minor, but common inhabitant of the gastrointestinal tract and opportunistic pathogen, including of the urinary tract, where it is the primary pathogen. While multi-omic analyses have detailed critical interactions between uropathogenic Escherichia coli (UPEC) and the bladder that mediate UTI outcome, comparatively little is known about UPEC in its pre-infection reservoir, partly due to its low abundance there (<1% relative abundance). To accurately and sensitively explore the genomes and transcriptomes of diverse E. coli in gastrointestinal communities, we developed E. coli PanSelect which uses a set of probes designed to specifically recognize and capture E. coli's broad pangenome from sequencing libraries. We demonstrated the ability of E. coli PanSelect to enrich, by orders of magnitude, sequencing data from diverse E. coli using a mock community and a set of human stool samples collected as part of a cohort study investigating drivers of recurrent urinary tract infections (rUTI). Comparisons of genomes and transcriptomes between E. coli residing in the gastrointestinal tracts of women with and without a history of rUTI suggest that rUTI gut E. coli are responding to increased levels of oxygen and nitrate, suggestive of mucosal inflammation, which may have implications for recurrent disease. E. coli PanSelect is well suited for investigations of native in vivo biology of E. coli in other environments where it is at low relative abundance, and the framework described here has broad applicability to other highly diverse, low abundance organisms.

Case report: Long-term follow-up of patients who received a FimCH vaccine for prevention of recurrent urinary tract infections caused by antibiotic resistant Enterobacteriaceae: a case report series.

Urinary tract infections (UTI) caused by carbapenem-resistant Enterobacteriaceae (CRE) are considered one of the most urgent health threats to humans according to the Centers for Disease Control (CDC), and the World Health Organization (WHO). A FimCH Vaccine expanded access study is being conducted in patients with a history of antibiotic resistant UTIs who are considered to be at risk for development of CRE UTI. This case series describes the clinical, safety and immunogenicity findings for four participants who received a FimCH four-vaccine series. Participants were followed for 12 months after administration of the fourth vaccine for safety, general health status and UTI occurrence. The study was later amended to allow additional follow-up of up to five years post vaccine administration to assess long-term health status, UTI occurrences and to obtain blood samples for anti-FimH antibody testing. In our population of 4 study participants, the number of symptomatic UTI occurrences caused by gram-negative bacteria in the 12-month period following peak anti-FimH antibody response were approximately 75% lower than the 12-month period preceding study enrollment. These results are consistent with the 30-patient cohort of a Phase 1 study with the same FimCH Vaccine. UTI occurrences increased during the long-term follow-up period for all 4 participants but did not reach the rate observed pre-vaccination. No new safety concerns related to the FimCH Vaccine were identified during long-term follow-up. This case series has clinical importance and public health relevance since it examines and reports on UTI frequency and recurrence following vaccination with the FimCH Vaccine in a high-risk population of patients with recurrent UTI. Additionally, participants described improved well-being following vaccination which was maintained in the long-term follow-up period.

Fibrinolytic-deficiencies predispose hosts to septicemia from a catheter-associated UTI.

Catheter-associated urinary tract infections (CAUTIs) are amongst the most common nosocomial infections worldwide and are difficult to treat partly due to development of multidrug-resistance from CAUTI-related pathogens. Importantly, CAUTI often leads to secondary bloodstream infections and death. A major challenge is to predict when patients will develop CAUTIs and which populations are at-risk for bloodstream infections. Catheter-induced inflammation promotes fibrinogen (Fg) and fibrin accumulation in the bladder which are exploited as a biofilm formation platform by CAUTI pathogens. Using our established mouse model of CAUTI, here we identified that host populations exhibiting either genetic or acquired fibrinolytic-deficiencies, inducing fibrin deposition in the catheterized bladder, are predisposed to severe CAUTI and septicemia by diverse uropathogens in mono- and poly-microbial infections. Furthermore, here we found that Enterococcus faecalis, a prevalent CAUTI pathogen, uses the secreted protease, SprE, to induce fibrin accumulation and create a niche ideal for growth, biofilm formation, and persistence during CAUTI.

Discovery of Orally Bioavailable FmlH Lectin Antagonists as Treatment for Urinary Tract Infections.

FmlH, a bacterial adhesin of uropathogenic Escherichia coli (UPEC), has been shown to provide a fitness advantage in colonizing the bladder during chronic urinary tract infections (UTIs). Previously reported ortho-biphenyl glycosides based on ßGal and ßGalNAc have excellent binding affinity to FmlH and potently block binding to its natural carbohydrate receptor, but they lack oral bioavailability. In this paper, we outline studies where we have optimized compounds for improved pharmacokinetics, leading to the discovery of novel analogues with good oral bioavailability. We synthesized galactosides with the anomeric O-linker replaced with more stable S- and C-linked linkers. We also investigated modifications to the GalNAc sugar and modifications to the biphenyl aglycone. We identified GalNAc 69 with an IC50 of 0.19 µM against FmlH and 53% oral bioavailability in mice. We also obtained a FimlH-bound X-ray structure of lead compound 69 (AM4085) which has potential as a new antivirulence therapeutic for UTIs.

Dihydrothiazolo ring-fused 2-pyridone antimicrobial compounds treat Streptococcus pyogenes skin and soft tissue infection.

We have developed GmPcides from a peptidomimetic dihydrothiazolo ring-fused 2-pyridone scaffold that have antimicrobial activities against a broad-spectrum of Gram-positive pathogens. Here we examine the treatment efficacy of GmPcides using skin and soft tissue infection (SSTI) and biofilm formation models by Streptococcus pyogenes. Screening our compound library for minimal inhibitory (MIC) and minimal bactericidal (MBC) concentrations identified GmPcide PS757 as highly active against S. pyogenes. Treatment of S. pyogenes biofilm with PS757 revealed robust efficacy against all phases of biofilm formation by preventing initial biofilm development, ceasing biofilm maturation and eradicating mature biofilm. In a murine model of S. pyogenes SSTI, subcutaneous delivery of PS757 resulted in reduced levels of tissue damage, decreased bacterial burdens and accelerated rates of wound-healing, which were associated with down-regulation of key virulence factors, including M protein and the SpeB cysteine protease. These data demonstrate that GmPcides show considerable promise for treating S. pyogenes infections.

Secretory leukocyte protease inhibitor protects against severe urinary tract infection in mice.

Millions suffer from urinary tract infections (UTIs) worldwide every year with women accounting for the majority of cases. Uropathogenic Escherichia coli (UPEC) causes most of these primary infections and leads to 25% becoming recurrent or chronic. To repel invading pathogens, the urinary tract mounts a vigorous innate immune response that includes the secretion of antimicrobial peptides (AMPs), rapid recruitment of phagocytes, and exfoliation of superficial umbrella cells. Here, we investigate secretory leukocyte protease inhibitor (SLPI), an AMP with antiprotease, antimicrobial, and immunomodulatory functions, known to play protective roles at other mucosal sites, but not well characterized in UTIs. Using a preclinical model of UPEC-caused UTI, we show that urine SLPI increases in infected mice and that SLPI is localized to bladder epithelial cells. UPEC-infected SLPI-deficient (Slpi-/-) mice suffer from higher urine bacterial burdens, prolonged bladder inflammation, and elevated urine neutrophil elastase (NE) levels compared to wild-type (Slpi+/+) controls. Combined with bulk bladder RNA sequencing, our data indicate that Slpi-/- mice have a dysregulated immune and tissue repair response following UTI. We also measure SLPI in urine samples from a small group of female subjects 18-49 years old and find that SLPI tends to be higher in the presence of a uropathogen, except in patients with a history of recent or recurrent UTI, suggesting a dysregulation of SLPI expression in these women. Taken together, our findings show SLPI promotes clearance of UPEC in mice and provides preliminary evidence that SLPI is likewise regulated in response to uropathogen exposure in women.IMPORTANCEAnnually, millions of people suffer from urinary tract infections (UTIs) and more than $3 billion are spent on work absences and treatment of these patients. While the early response to UTI is known to be important in combating urinary pathogens, knowledge of host factors that help curb infection is still limited. Here, we use a preclinical model of UTI to study secretory leukocyte protease inhibitor (SLPI), an antimicrobial protein, to determine how it protects the bladder against infection. We find that SLPI is increased during UTI, accelerates the clearance of bacteriuria, and upregulates genes and pathways needed to fight an infection while preventing prolonged bladder inflammation. In a small clinical study, we show SLPI is readily detectable in human urine and is associated with the presence of a uropathogen in patients without a previous history of UTI, suggesting SLPI may play an important role in protecting from bacterial cystitis.

Microbial co-occurrences on catheters from long-term catheterized patients.

Catheter-associated urinary tract infections (CAUTIs), a common cause of healthcare-associated infections, are caused by a diverse array of pathogens that are increasingly becoming antibiotic resistant. We analyze the microbial occurrences in catheter and urine samples from 55 human long-term catheterized patients collected over one year. Although most of these patients were prescribed antibiotics over several collection periods, their catheter samples remain colonized by one or more bacterial species. Examination of a total of 366 catheter and urine samples identify 13 positive and 13 negative genus co-occurrences over 12 collection periods, representing associations that occur more or less frequently than expected by chance. We find that for many patients, the microbial species composition between collection periods is similar. In a subset of patients, we find that the most frequently sampled bacteria, Escherichia coli and Enterococcus faecalis, co-localize on catheter samples. Further, co-culture of paired isolates recovered from the same patients reveals that E. coli significantly augments E. faecalis growth in an artificial urine medium, where E. faecalis monoculture grows poorly. These findings suggest novel strategies to collapse polymicrobial CAUTI in long-term catheterized patients by targeting mechanisms that promote positive co-associations.

Secretory Leukocyte Protease Inhibitor Protects Against Severe Urinary Tract Infection in Mice.

Millions suffer from urinary tract infections (UTIs) worldwide every year with women accounting for the majority of cases. Uropathogenic Escherichia coli (UPEC) causes most of these primary infections and leads to 25% becoming recurrent or chronic. To repel invading pathogens, the urinary tract mounts a vigorous innate immune response that includes the secretion of antimicrobial peptides (AMPs), rapid recruitment of phagocytes and exfoliation of superficial umbrella cells. Here, we investigate secretory leukocyte protease inhibitor (SLPI), an AMP with antiprotease, antimicrobial and immunomodulatory functions, known to play protective roles at other mucosal sites, but not well characterized in UTIs. Using a mouse model of UPEC-caused UTI, we show that urine SLPI increases in infected mice and that SLPI is localized to bladder epithelial cells. UPEC infected SLPI-deficient (Slpi-/-) mice suffer from higher urine bacterial burdens, prolonged bladder inflammation, and elevated urine neutrophil elastase (NE) levels compared to wild-type (Slpi+/+) controls. Combined with bulk bladder RNA sequencing, our data indicate that Slpi-/- mice have a dysregulated immune and tissue repair response following UTI. We also measure SLPI in urine samples from a small group of female subjects 18-49 years old and find that SLPI tends to be higher in the presence of a uropathogen, except in patients with history of recent or recurrent UTI (rUTI), suggesting a dysregulation of SLPI expression in these women. Taken together, our findings show SLPI protects against acute UTI in mice and provides preliminary evidence that SLPI is likewise regulated in response to uropathogen exposure in women.

Fibrinolytic-deficiencies predispose hosts to septicemia from a catheter-associated UTI.

Catheter-associated urinary tract infections (CAUTIs) are amongst the most common nosocomial infections worldwide and are difficult to treat due to multi-drug resistance development among the CAUTI-related pathogens. Importantly, CAUTI often leads to secondary bloodstream infections and death. A major challenge is to predict when patients will develop CAUTIs and which populations are at-risk for bloodstream infections. Catheter-induced inflammation promotes fibrinogen (Fg) and fibrin accumulation in the bladder which are exploited as a biofilm formation platform by CAUTI pathogens. Using our established mouse model of CAUTI, we identified that host populations exhibiting either genetic or acquired fibrinolytic-deficiencies, inducing fibrin deposition in the catheterized bladder, are predisposed to severe CAUTI and septicemia by diverse uropathogens in mono- and poly-microbial infections. Furthermore, we found that E. faecalis, a prevalent CAUTI pathogen, uses the secreted protease, SprE, to induce fibrin accumulation and create a niche ideal for growth, biofilm formation, and persistence during CAUTI.

A prospective randomized clinical trial to assess antibiotic pocket irrigation on tissue expander breast reconstruction.

Bacterial infection is the most common complication following staged post-mastectomy breast reconstruction initiated with a tissue expander (TE). To limit bacterial infection, antibiotic irrigation of the surgical site is commonly performed despite little high-quality data to support this practice. We performed a prospective randomized control trial to compare the impact of saline irrigation alone to a triple antibiotic irrigation regimen (1 g cefazolin, 80 mg gentamicin, and 50,000 units of bacitracin in 500 mL of saline) for breast implant surgery. The microbiome in breasts with cancer (n = 16) was compared to those without (n = 16), as all patients (n = 16) had unilateral cancers but bilateral mastectomies (n = 32). Biologic and prosthetic specimens procured both at the time of mastectomy and during TE removal months later were analyzed for longitudinal comparison. Outcomes included clinical infection, bacterial abundance, and relative microbiome composition. No patient in either group suffered a reconstructive failure or developed an infection. Triple antibiotic irrigation administered at the time of immediate TE reconstruction did not reduce bacterial abundance or impact microbial diversity relative to saline irrigation at the time of planned exchange. Implanted prosthetic material adopted the microbial composition of the surrounding host tissue. In cancer-naïve breasts, relative to saline, antibiotic irrigation increased bacterial abundance on periprosthetic capsules (P = 0.03) and acellular dermal matrices (P = 0.04) and altered the microbiota on both. These data show that, relative to saline only, the use of triple antibiotic irrigation in TE breast reconstruction does impact the bacterial abundance and diversity of certain biomaterials from cancer-naïve breasts. IMPORTANCE The lifetime risk of breast cancer is ~13% in women and is treated with a mastectomy in ~50% of cases. The majority are reconstructed, usually starting with a tissue expander to help restore the volume for a subsequent permanent breast implant or the women's own tissues. The biopsychosocial benefits of breast reconstruction, though, can be tempered by a high complication rate of at least 7% but over 30% in some women. Bacterial infection is the most common complication, and can lead to treatment delays, patient physical and emotional distress and escalating health care cost. To limit this risk, plastic surgeons have tried a variety of strategies to limit bacterial infection including irrigating the pocket created after removing the breast implant with antibiotic solutions, but good-quality data are scarce. Herein, we study the value of antibiotics in pocket irrigation using a robust randomized clinical trial design and molecular microbiology approaches.

Vaccination with Acinetobacter baumannii adhesin Abp2D provides protection against catheter-associated urinary tract infection.

Catheter-associated urinary tract infections (CAUTIs) contribute greatly to the burden of healthcare associated infections. Acinetobacter baumannii is a Gram-negative bacterium with high levels of antibiotic resistance that is of increasing concern as a CAUTI pathogen. A. baumannii expresses fibrinogen-binding adhesins (Abp1D and Abp2D) that mediate colonization and biofilm formation on catheters, which become coated with fibrinogen upon insertion. We developed a protein subunit vaccine against Abp1DRBD and Abp2DRBD and showed that vaccination significantly reduced bladder bacterial titers in a mouse model of CAUTI. We then determined that immunity to Abp2DRBD alone was sufficient for protection. Mechanistically, we defined the B cell response to Abp2DRBD vaccination and demonstrated that immunity was transferrable to naïve mice through passive immunization with Abp2DRBD-immune sera. This work represents a novel strategy in the prevention of A. baumannii CAUTI and has an important role to play in the global fight against antimicrobial resistance.

Defining the Roles of Pyruvate Oxidation, TCA Cycle, and Mannitol Metabolism in Methicillin-Resistant Staphylococcus aureus Catheter-Associated Urinary Tract Infection.

Methicillin-resistant Staphylococcus aureus (MRSA) is an important cause of complicated urinary tract infection (UTI) associated with the use of indwelling urinary catheters. Previous reports have revealed host and pathogen effectors critical for MRSA uropathogenesis. Here, we sought to determine the significance of specific metabolic pathways during MRSA UTI. First, we identified four mutants from the Nebraska transposon mutant library in the MRSA JE2 background that grew normally in rich medium but displayed significantly reduced growth in pooled human urine (HU). This prompted us to transduce the uropathogenic MRSA 1369 strain with the transposon mutants in sucD and fumC (tricarboxylic acid [TCA] cycle), mtlD (mannitol metabolism), and lpdA (pyruvate oxidation). Notably, sucD, fumC, and mtlD were also significantly upregulated in the MRSA 1369 strain upon exposure to HU. Compared to the WT, the MRSA 1369 lpdA mutant was significantly defective for (i) growth in HU, and (ii) colonization of the urinary tract and dissemination to the kidneys and the spleen in the mouse model of catheter-associated UTI (CAUTI), which may be attributed to its increased membrane hydrophobicity and higher susceptibility to killing by human blood. In contrast to their counterparts in the JE2 background, the sucD, fumC, and mtlD mutants in the MRSA 1369 background grew normally in HU; however, they displayed significant fitness defects in the CAUTI mouse model. Overall, identification of novel metabolic pathways important for the urinary fitness and survival of MRSA can be used for the development of novel therapeutics. IMPORTANCE While Staphylococcus aureus has historically not been considered a uropathogen, S. aureus urinary tract infection (UTI) is clinically significant in certain patient populations, including those with chronic indwelling urinary catheters. Moreover, most S. aureus strains causing catheter-associated UTI (CAUTI) are methicillin-resistant S. aureus (MRSA). MRSA is difficult to treat due to limited treatment options and the potential to deteriorate into life-threatening bacteremia, urosepsis, and shock. In this study, we found that pathways involved in pyruvate oxidation, TCA cycle, and mannitol metabolism are important for MRSA fitness and survival in the urinary tract. Improved understanding of the metabolic needs of MRSA in the urinary tract may help us develop novel inhibitors of MRSA metabolism that can be used to treat MRSA-CAUTI more effectively.

Uropathogenic Escherichia coli infection-induced epithelial trained immunity impacts urinary tract disease outcome.

Previous urinary tract infections (UTIs) can predispose one to future infections; however, the underlying mechanisms affecting recurrence are poorly understood. We previously found that UTIs in mice cause differential bladder epithelial (urothelial) remodelling, depending on disease outcome, that impacts susceptibility to recurrent UTI. Here we compared urothelial stem cell (USC) lines isolated from mice with a history of either resolved or chronic uropathogenic Escherichia coli (UPEC) infection, elucidating evidence of molecular imprinting that involved epigenetic changes, including differences in chromatin accessibility, DNA methylation and histone modification. Epigenetic marks in USCs from chronically infected mice enhanced caspase-1-mediated cell death upon UPEC infection, promoting bacterial clearance. Increased Ptgs2os2 expression also occurred, potentially contributing to sustained cyclooxygenase-2 expression, bladder inflammation and mucosal wounding-responses associated with severe recurrent cystitis. Thus, UPEC infection acts as an epi-mutagen reprogramming the urothelial epigenome, leading to urothelial-intrinsic remodelling and training of the innate response to subsequent infection.

Urinary tract infections trigger synucleinopathy via the innate immune response.

Symptoms in the urogenital organs are common in multiple system atrophy (MSA), also in the years preceding the MSA diagnosis. It is unknown how MSA is triggered and these observations in prodromal MSA led us to hypothesize that synucleinopathy could be triggered by infection of the genitourinary tract causing ɑ-synuclein (ɑSyn) to aggregate in peripheral nerves innervating these organs. As a first proof that peripheral infections could act as a trigger in MSA, this study focused on lower urinary tract infections (UTIs), given the relevance and high frequency of UTIs in prodromal MSA, although other types of infection might also be important triggers of MSA. We performed an epidemiological nested-case control study in the Danish population showing that UTIs are associated with future diagnosis of MSA several years after infection and that it impacts risk in both men and women. Bacterial infection of the urinary bladder triggers synucleinopathy in mice and we propose a novel role of ɑSyn in the innate immune system response to bacteria. Urinary tract infection with uropathogenic E. coli results in the de novo aggregation of ɑSyn during neutrophil infiltration. During the infection, ɑSyn is released extracellularly from neutrophils as part of their extracellular traps. Injection of MSA aggregates into the urinary bladder leads to motor deficits and propagation of ɑSyn pathology to the central nervous system in mice overexpressing oligodendroglial ɑSyn. Repeated UTIs lead to progressive development of synucleinopathy with oligodendroglial involvement in vivo. Our results link bacterial infections with synucleinopathy and show that a host response to environmental triggers can result in ɑSyn pathology that bears semblance to MSA.

Strain Tracking with Uncertainty Quantification.

The ability to detect and quantify microbiota over time has a plethora of clinical, basic science, and public health applications. One of the primary means of tracking microbiota is through sequencing technologies. When the microorganism of interest is well characterized or known a priori, targeted sequencing is often used. In many applications, however, untargeted bulk (shotgun) sequencing is more appropriate; for instance, the tracking of infection transmission events and nucleotide variants across multiple genomic loci, or studying the role of multiple genes in a particular phenotype. Given these applications, and the observation that pathogens (e.g. Clostridioides difficile, Escherichia coli, Salmonella enterica) and other taxa of interest can reside at low relative abundance in the gastrointestinal tract, there is a critical need for algorithms that accurately track low-abundance taxa with strain level resolution. Here we present a sequence quality- and time-aware model, ChronoStrain, that introduces uncertainty quantification to gauge low-abundance species and significantly outperforms the current state-of-the-art on both real and synthetic data. ChronoStrain leverages sequences' quality scores and the samples' temporal information to produce a probability distribution over abundance trajectories for each strain tracked in the model. We demonstrate Chronostrain's improved performance in capturing post-antibiotic E. coli strain blooms among women with recurrent urinary tract infections (UTIs) from the UTI Microbiome (UMB) Project. Other strain tracking models on the same data either show inconsistent temporal colonization or can only track consistently using very coarse groupings. In contrast, our probabilistic outputs can reveal the relationship between low-confidence strains present in the sample that cannot be reliably assigned a single reference label (either due to poor coverage or novelty) while simultaneously calling high-confidence strains that can be unambiguously assigned a label. We also include and analyze newly sequenced cultured samples from the UMB Project.

Catheterization of mice triggers resurgent urinary tract infection seeded by a bladder reservoir of Acinetobacter baumannii.

The antibiotic-resistant bacterium Acinetobacter baumannii is a leading cause of hospital-associated infections. Despite surveillance and infection control efforts, new A. baumannii strains are regularly isolated from health care facilities worldwide. In a mouse model of urinary tract infection, we found that mice infected with A. baumannii displayed high bacterial burdens in urine for several weeks. Two months after the resolution of A. baumannii infection, inserting a catheter into the bladder of mice with resolved infection led to the resurgence of a same-strain urinary tract infection in ~53% of the mice within 24 hours. We identified intracellular A. baumannii bacteria in the bladder epithelial cells of mice with resolved infection, which we propose could act as a host reservoir that was activated upon insertion of a catheter, leading to a resurgent secondary infection.

Structure-function correlates of fibrinogen binding by Acinetobacter adhesins critical in catheter-associated urinary tract infections.

Multidrug-resistant Acinetobacter baumannii infections are an urgent clinical problem and can cause difficult-to-treat nosocomial infections. During such infections, like catheter-associated urinary tract infections (CAUTI), A. baumannii rely on adhesive, extracellular fibers, called chaperone-usher pathway (CUP) pili for critical binding interactions. The A. baumannii uropathogenic strain, UPAB1, and the pan-European subclone II isolate, ACICU, use the CUP pili Abp1 and Abp2 (previously termed Cup and Prp, respectively) in tandem to establish CAUTIs, specifically to facilitate bacterial adherence and biofilm formation on the implanted catheter. Abp1 and Abp2 pili are tipped with two domain tip adhesins, Abp1D and Abp2D, respectively. We discovered that both adhesins bind fibrinogen, a critical host wound response protein that is released into the bladder upon catheterization and is subsequently deposited on the catheter. The crystal structures of the Abp1D and Abp2D receptor-binding domains were determined and revealed that they both contain a large, distally oriented pocket, which mediates binding to fibrinogen and other glycoproteins. Genetic, biochemical, and biophysical studies revealed that interactions with host proteins are governed by several critical residues in and along the edge of the binding pocket, one of which regulates the structural stability of an anterior loop motif. K34, located outside of the pocket but interacting with the anterior loop, also regulates the binding affinity of the protein. This study illuminates the mechanistic basis of the critical fibrinogen-coated catheter colonization step in A. baumannii CAUTI pathogenesis.

Ring-fused 2-pyridones effective against multidrug-resistant Gram-positive pathogens and synergistic with standard-of-care antibiotics.

The alarming rise of multidrug-resistant Gram-positive bacteria has precipitated a healthcare crisis, necessitating the development of new antimicrobial therapies. Here we describe a new class of antibiotics based on a ring-fused 2-pyridone backbone, which are active against vancomycin-resistant enterococci (VRE), a serious threat as classified by the Centers for Disease Control and Prevention, and other multidrug-resistant Gram-positive bacteria. Ring-fused 2-pyridone antibiotics have bacteriostatic activity against actively dividing exponential phase enterococcal cells and bactericidal activity against nondividing stationary phase enterococcal cells. The molecular mechanism of drug-induced killing of stationary phase cells mimics aspects of fratricide observed in enterococcal biofilms, where both are mediated by the Atn autolysin and the GelE protease. In addition, combinations of sublethal concentrations of ring-fused 2-pyridones and standard-of-care antibiotics, such as vancomycin, were found to synergize to kill clinical strains of VRE. Furthermore, a broad range of antibiotic resistant Gram-positive pathogens, including those responsible for the increasing incidence of antibiotic resistant healthcare-associated infections, are susceptible to this new class of 2-pyridone antibiotics. Given the broad antibacterial activities of ring-fused 2-pyridone compounds against Gram-positive (GmP) bacteria we term these compounds GmPcides, which hold promise in combating the rising tide of antibiotic resistant Gram-positive pathogens.

Complete Genome Sequence of the Uropathogenic Methicillin-Resistant Staphylococcus aureus Strain MRSA-1369.

MRSA-1369 is a uropathogenic methicillin-resistant Staphylococcus aureus (MRSA) strain. Here, we present the complete genome sequence of MRSA-1369, which consists of one chromosome (2.87 Mb) and two plasmids (16.68 kb and 3.13 kb). This will serve as a reference genome for future Staphylococcus aureus pathogenesis and multiomic studies.

Characterization of Host-Pathogen-Device Interactions in Pseudomonas aeruginosa Infection of Breast Implants.

BACKGROUND: Pseudomonas aeruginosa accounts for 7 to 22 percent of breast implant-associated infections, which can result in reconstructive failures and explantation. Investigating host-pathogen-device interactions in mice and patient samples will improve the understanding of colonization mechanisms, for targeted treatments and clinical guidelines. METHODS: Mice with and without implants were infected with PAO1 laboratory strain or BIP2 or BIP16 clinical strains and killed at 1 day or 7 days after infection to evaluate for colonization of implants and underlying tissues by means of colony-forming unit enumeration. Immunostaining was performed on mouse implants, human tissue expanders colonized by BIP2, and acellular dermal matrix colonized by BIP16. RESULTS: Colonization of tissues and smooth implants by P. aeruginosa was strain-dependent: at 1 day after infection, all strains acutely infected tissues with and without implants with colonization levels reflecting growth rates of individual strains. At 7 days after infection, PAO1 caused colonization of approximately 10 5 colony-forming units/100 mg of tissue but required implant presence, whereas in mice infected with BIP2/BIP16, colony-forming units were below the limit of detection with or without implants. Immunofluorescence staining of mouse implants, however, demonstrated continued presence of BIP2 and BIP16. Staining showed co-localization of all strains with fibrinogen, collagen I, and collagen III on mouse and human samples. CONCLUSIONS: The trajectory of P. aeruginosa in breast implant-associated infections was strain-dependent, and strains could exhibit acute symptomatic or chronic asymptomatic colonization. With strains causing clinical symptoms, the presence of an implant significantly worsened infection. For asymptomatic colonizers, further studies investigating their long-term impacts, especially during periods of immunosuppression in hosts, are needed.