IMPORTANCE:Urinary tract infections (UTIs) are common in older-aged women. Our study examined bacterial persistence with commonly prescribed antibiotics. Bacterial growth was demonstrated despite antibiotic treatment. OBJECTIVES:The aims of this study were to quantify the bacterial persister phenotype in urine collected from postmenopausal women with acute and recurrent UTI and to determine the capabilities of first-line antibiotics to effectively treat persister cells. STUDY DESIGN:This was an institutional review board-approved cross-sectional analysis within a large academic referral center. Uropathogens were cultured from postmenopausal women with acute or recurrent UTI and screened for persister cells using persistence assays. Demographic and clinical variables were collected and analyzed. The entire experimental process was repeated in triplicate. Data were analyzed for significance (P < 0.05) between the persister culture and antibiotic treatments using a 1-way analysis of variance with multiple comparisons in Prism 9.3.0. RESULTS:Forty participants were included: 62.5% White, 22.5% Black, 3% Asian, and 2% Hispanic with a mean age of 72.3 ± 11.62 years. The persister phenotype was demonstrated in all of Escherichia coli isolates. Treatment with fosfomycin demonstrated reduced colony-forming units per milliliter compared with control (P < 0.01). Among recurrent isolates, there was a statistically significant decrease in colony-forming units per milliliter after antibiotic treatment with all 4 antibiotics (P < 0.05). CONCLUSIONS:This study demonstrated in vitro bacterial persistence in uropathogens from urogynecology patients despite treatment with commonly prescribed antibiotics. Fosfomycin generated the least amount of persister cells. Results suggest that persistence may be one bacterial defense mechanism involved in UTIs. Further research is needed to understand the clinical implications.

Quaternary ammonium compounds (QACs) serve as a first line of defense against infectious pathogens. As resistance to QACs emerges in the environment, the development of next-generation disinfectants is of utmost priority for human health. Balancing antibacterial potency with environmental considerations is required to effectively counter the development of bacterial resistance. To address this challenge, a series of 14 novel biscationic quaternary phosphonium compounds (bisQPCs) have been prepared as amphiphilic disinfectants through straightforward, high-yielding alkylation reactions. These compounds feature decomposable or "soft" amide moieties in their side chains, anticipated to promote decomposition under environmental conditions. Strong bioactivity against a panel of seven bacterial pathogens was observed, highlighted by single-digit micromolar activity for compounds P6P-12A,12A and P3P-12A,12A. Hydrolysis experiments in pure water and in buffers of varying pH revealed surprising decomposition of the soft QPCs under basic conditions at the phosphonium center, leading to inactive phosphine oxide products; QPC stability (>24 h) was maintained in neutral solutions. The results of this work unveil soft QPCs as a potent and environmentally conscious new class of bisQPC disinfectants.

Throughout the SARS-CoV-2 pandemic, the use of botanical dietary supplements in the United States has increased, yet their safety and efficacy against COVID-19 remains underexplored. The Quave Natural Product Library is a phylogenetically diverse collection of botanical and fungal natural product extracts including popular supplement ingredients. Evaluation of 1867 extracts and 18 compounds for virus spike protein binding to host cell ACE2 receptors in a SARS-CoV-2 pseudotyped virus system identified 310 extracts derived from 188 species across 76 families (3 fungi, 73 plants) that exhibited ≥ 50% viral entry inhibition activity at 20 µg/mL. Extracts exhibiting mammalian cytotoxicity > 15% and those containing cardiotoxic cardiac glycosides were eliminated. Three extracts were selected for further testing against four pseudotyped variants and infectious SARS-CoV-2 and were then further chemically characterized, revealing the potent (EC 50 < 5 µg/mL) antiviral activity of Solidago altissima L. (Asteraceae) flowers and Pteridium aquilinum (L.) Kuhn (Dennstaedtiaceae) rhizomes.

Covering: 2018 to 2022Antimicrobial resistance (AMR) poses a significant global health threat. There is a rising demand for innovative drug scaffolds and new targets to combat multidrug-resistant bacteria. Before the advent of antibiotics, infections were treated with plants chosen from traditional medicine practices. Of Earth's 374 000 plant species, approximately 9% have been used medicinally, but most species remain to be investigated. This review illuminates discoveries of antimicrobial natural products from plants covering 2018 to 2022. It highlights plant-derived natural products with antibacterial, antivirulence, and antibiofilm activity documented in lab studies. Additionally, this review examines the development of novel derivatives from well-studied parent natural products, as natural product derivatives have often served as scaffolds for anti-infective agents.

The examination of biodiversity across the world has historically been a critical part of drug development and led to the discovery of common medications for many medical issues including pain management, cancer, heart disease, and infections. During the SARS‐CoV‐2 pandemic, the use of natural supplements in the United States has increased. The efficacy of these natural products to prevent SARS‐CoV‐2 infection and the safety of their use remains unexplored; therefore, more research must be done to determine which supplements have antiviral properties. The Quave Natural Product Library (QNPL) is a collection of over 2,000 botanical and fungal extracts and includes the 40 most used natural supplements in the United States. Collection of the biological samples for the library requires field expeditions to areas throughout the world with high levels of biodiversity. Each of these extracts were tested in a SARS‐CoV‐2 pseudotyped virus system to determine which extracts inhibit viral entry, specifically the virus spike protein binding to host cells ACE2 receptors. Mammalian cell cytotoxicity assays were run in parallel. Evaluation of 1,887 extracts and 18 single compounds from the QNPL against SARS‐CoV‐2 identified. 317 extracts derived from 134 species across 76 families (1 lichen, 2 fungi, 73 plant families) exhibited ≥50% inhibition activity in the wild type spike pseudotyped model at 20 µg/mL. Of these bioactive extracts, 129 extracts derived from 95 plant species exhibited ≥85% inhibition activity and ≤15% cytotoxicity in the wild‐type model. Once these 129 extracts were identified, an interesting pattern emerged indicating many hits were from species that are known to be cardiotoxic due to rich composition of cardiac glycosides. For further selection and testing, we reviewed each extract and consulted the literature to eliminate extracts with those properties or similar compounds, which narrowed down our interest to 8 extracts. These extracts were further validated in a concentration‐response assay in a pseudotyped virus model. The EC50 values of the top 3 extracts were all under 10 µg/mL. These 3 extracts all exhibited activity (≥85% inhibition activity) in the wildtype and variant pseudotyped models. Testing in live SARS‐CoV‐2 confirmed antiviral activity from 2 of 3 extracts, Plant Aflowers and Plant B roots. Further chemical characterization of the major metabolites of these two hits was performed using MS/MS fragmentation data compared with the literature, in silico prediction, and web‐based databases. The results revealed phenylpropanoids, flavonoids, triterpenes, glycosidic terpenes, and fatty acids as the major chemical classes. The next steps of this study seek to identify and isolate purified bioactive compounds to further understand their role in SARS‐CoV‐2 inhibition.

Methicillin-resistant Staphylococcus aureus (MRSA) represents one of the most serious infectious disease concerns worldwide, with the CDC labeling it a "serious threat" in 2019. The current arsenal of antibiotics works by targeting bacterial growth and survival, which exerts great selective pressure for the development of resistance. The development of novel anti-infectives that inhibit quorum sensing and thus virulence in MRSA has been recurrently proposed as a promising therapeutic approach. In a follow-up of a study examining the MRSA quorum sensing inhibitory activity of extracts of Italian plants used in local traditional medicine, 224C-F2 was reported as a bioactive fraction of a Castanea sativa (European chestnut) leaf extract. The fraction demonstrated high activity in vitro and effective attenuation of MRSA pathogenicity in a mouse model of skin infection. Through further bioassay-guided fractionation using reverse-phase high performance liquid chromatography, a novel hydroperoxy cycloartane triterpenoid, castaneroxy A (1), was isolated. Its structure was established by nuclear magnetic resonance, mass spectrometry and X-ray diffraction analyses. Isomers of 1 were also detected in an adjacent fraction. In a series of assays assessing inhibition of markers of MRSA virulence, 1 exerted activities in the low micromolar range. It inhibited agr::P3 activation (IC50 = 31.72 µM), δ-toxin production (IC50 = 31.72 µM in NRS385), supernatant cytotoxicity to HaCaT human keratinocytes (IC50 = 7.93 µM in NRS385), and rabbit erythrocyte hemolytic activity (IC50 = 7.93 µM in LAC). Compound 1 did not inhibit biofilm production, and at high concentrations it exerted cytotoxicity against human keratinocytes greater than that of 224C-F2. Finally, 1 reduced dermonecrosis in a murine model of MRSA infection. The results establish 1 as a promising antivirulence candidate for development against MRSA.

The rise of antibiotic resistance presents a significant healthcare challenge and precludes the use of many otherwise valuable antibiotics. One potential solution to this problem is the use of antibiotics in combination with resistance-modifying agents, compounds that act synergistically with existing antibiotics to resensitize previously resistant bacteria. In this study, 12(S),16ξ-dihydroxycleroda-3,13-dien-15,16-olide, a clerodane diterpene isolated from the medicinal plant Callicarpa americana, was found to synergize with oxacillin against methicillin-resistant Staphylococcus aureus. This synergy was confirmed by checkerboard (FICI = 0.125) and time-kill assays, with a sub-inhibitory dose of 12(S),16ξ-dihydroxycleroda-3,13-dien-15,16-olide causing the effective concentration of oxacillin to fall below the susceptibility breakpoint for S. aureus, a > 32-fold decrease in both cases.

Calcium-mediated signaling through inositol 1,4,5-triphosphate receptors (IP3Rs) is essential for the regulation of numerous physiological processes, including fertilization, muscle contraction, apoptosis, secretion, and synaptic plasticity. Deregulation of IP3Rs leads to pathological calcium signaling and is implicated in many common diseases, including cancer and neurodegenerative, autoimmune, and metabolic diseases. Revealing the mechanism of activation and inhibition of this ion channel will be critical to an improved understanding of the biological processes that are controlled by IP3Rs. Here, we report structural findings of the human type-3 IP3R (IP3R-3) obtained by cryo-EM (at an overall resolution of 3.8 Å), revealing an unanticipated regulatory mechanism where a loop distantly located in the primary sequence occupies the IP3-binding site and competitively inhibits IP3 binding. We propose that this inhibitory mechanism must differ qualitatively among IP3R subtypes because of their diverse loop sequences, potentially serving as a key molecular determinant of subtype-specific calcium signaling in IP3Rs. In summary, our structural characterization of human IP3R-3 provides critical insights into the mechanistic function of IP3Rs and into subtype-specific regulation of these important calcium-regulatory channels.

Atopic dermatitis (AD) represents a complex intersection between skin barrier dysfunction, host immunologic response, and external factors such as allergenic triggers and the skin microbiome. However, the relative contributions of these factors are not clear. There is a surge in Staphylococcus aureus on the skin during flares of AD (Kong et al., 2012Kong H.H. Oh J. Deming C. Conlan S. Grice E.A. Beatson M.A. et al.Temporal shifts in the skin microbiome associated with disease flares and treatment in children with atopic dermatitis.Genome Res. 2012; 22: 850-859Crossref PubMed Scopus (998) Google Scholar). S. aureus strains seen in AD flares are unique to the host and capable of eliciting varied immune responses (Byrd et al., 2017Byrd A.L. Deming C. Cassidy S.K.B. Harrison O.J. Ng W.I. Conlan S. et al.Staphylococcus aureus and Staphylococcus epidermidis strain diversity underlying pediatric atopic dermatitis.Sci Transl Med. 2017; 9eaal4651Crossref PubMed Scopus (244) Google Scholar). No direct relationship between severity and in vitro toxin genotype has been established (Kim et al., 2009Kim D.W. Park J.Y. Park K.D. Kim T.H. Lee W.J. Lee S.J. et al.Are there predominant strains and toxins of Staphylococcus aureus in atopic dermatitis patients? Genotypic characterization and toxin determination of S. aureus isolated in adolescent and adult patients with atopic dermatitis.J Dermatol. 2009; 36: 75-81Crossref PubMed Scopus (36) Google Scholar). S. epidermidis may similarly contribute to the propagation of AD through the elucidation of toxins (Martínez-García et al., 2018Martínez-García S. Rodríguez-Martínez S. Cancino-Diaz M.E. Cancino-Diaz J.C. Extracellular proteases of Staphylococcus epidermidis: roles as virulence factors and their participation in biofilm.APMIS. 2018; 126: 177-185Crossref PubMed Scopus (27) Google Scholar). Further information on strains of staphylococci found on patients with AD may aid in developing therapeutic targets. δ-Toxin (also known as δ-hemolysin) is produced by 97% of S. aureus strains as well as many coagulase-negative staphylococci (Dinges et al., 2000Dinges M.M. Orwin P.M. Schlievert P.M. Exotoxins of Staphylococcus aureus.Clin Microbiol Rev. 2000; 13: 16-34Crossref PubMed Scopus (1260) Google Scholar). Staphylococcal δ-toxin lyses mammalian erythrocytes and is particularly potent against rabbit erythrocytes (Kreger et al., 1971Kreger A.S. Kim K.S. Zaboretzky F. Bernheimer A.W. Purification and properties of staphylococcal delta hemolysin.Infect Immun. 1971; 3: 449-465Crossref PubMed Google Scholar); it also induces mast cell degranulation in vitro and is capable of inducing inflammatory skin disease in mouse models (Nakamura et al., 2013Nakamura Y. Oscherwitz J. Cease K.B. Chan S.M. Muñoz-Planillo R. Hasegawa M. et al.Staphylococcus δ-toxin induces allergic skin disease by activating mast cells.Nature. 2013; 503: 397-401Crossref PubMed Scopus (310) Google Scholar). Variability in δ-toxin production by S. aureus strains is influenced by complex interactions between a number of genes, with the carA gene playing a critical role in its production (Su et al., 2020Su M. Lyles J.T. Petit Iii Iii R.A. Peterson J. Hargita M. Tang H. et al.Genomic analysis of variability in delta-toxin levels between Staphylococcus aureus strains.PeerJ. 2020; 8: e8717Crossref PubMed Scopus (7) Google Scholar). In this study, we report data from a pilot study to begin to define the relationship between hemolytic toxin production and AD severity in pediatric patients and young adults. The Emory University (Atlanta, GA) Institutional Review Board approved this study (Institutional Review Board #0087896 and 00093739). Written informed consent and/or assent was obtained from parents or patients aged 2–20 years with AD. The diagnosis of AD was confirmed by a board-certified pediatric dermatologist after initial screening on the basis of the criteria outlined by Hanifin and Rajka, 1980Hanifin J.M. Rajka G. Diagnostic features of atopic dermatitis.Acta Derm Venereol. 1980; : 44-47Google Scholar. Patients were recruited from the Emory University pediatric dermatology clinics from May 2016 to May 2018 and were not asked to modify their topical or oral medications before this study to collect preliminary data on this subject. We assessed disease severity at the time of sample collection on the basis of the SCORing Atopic Dermatitis Index and grouped it into mild, moderate, or severe (Chopra et al., 2017Chopra R. Vakharia P.P. Sacotte R. Patel N. Immaneni S. White T. et al.Severity strata for Eczema Area and Severity Index (EASI), modified EASI, Scoring Atopic Dermatitis (SCORAD), objective SCORAD, Atopic Dermatitis Severity Index and body surface area in adolescents and adults with atopic dermatitis.Br J Dermatol. 2017; 177: 1316-1321Crossref PubMed Scopus (99) Google Scholar). Patients taking oral antibiotics at the time of the visit were excluded. Swabs were collected from the three most significantly flared body sites from areas of active dermatitis (affected), unaffected volar forearm without active dermatitis, and anterior nares. Swabs from the antecubital fossa, volar forearm, and nares were collected from nine healthy controls. Details of sample collection and processing are available in Supplementary Material and Methods S1. A total of 30 patients and/or parents provided written consent for participation in this study. Most patients had a moderate (n = 12) SCORing Atopic Dermatitis Index score, with fewer mild (n = 5) and severe (n = 4) scores (Chopra et al., 2017Chopra R. Vakharia P.P. Sacotte R. Patel N. Immaneni S. White T. et al.Severity strata for Eczema Area and Severity Index (EASI), modified EASI, Scoring Atopic Dermatitis (SCORAD), objective SCORAD, Atopic Dermatitis Severity Index and body surface area in adolescents and adults with atopic dermatitis.Br J Dermatol. 2017; 177: 1316-1321Crossref PubMed Scopus (99) Google Scholar) (Supplementary Table S1). Samples were collected from nine patients without AD. The skin of severely affected patients had the highest staphylococcal density in all pooled samples (Supplementary Table S2) as well as relative to the affected skin in patients with mild or moderate AD (Figure 1). Density was not significantly different in nares or unaffected skin between mild, moderate, and severe patients (Figure 1). Hemolytic potential of 1,054 isolates was assessed in a rabbit blood lysis model (Supplementary Figure S1). There was a significant increase in average hemolytic activity of staphylococcal isolates pooled from all collected sites from mild patients relative to those from moderate patients and those from moderate patients relative to those from severe patients (Figure 2a and Supplementary Table S3), whereas hemolysis in control isolates was not significantly different from that in mild patients. A similar stepwise increase in average hemolytic activity was demonstrated in isolates from unaffected skin of patients with AD, as well as from nares, and affected skin when analyzed independently (Figure 2b‒d). Body site and patient-level data are provided in Supplementary Figures S2 and S3. Although severe patients were most heavily colonized with staphylococci in affected skin (Supplementary Table S2), our study aligns with previous studies showing that staphylococcal load alone does not consistently correlate with the severity of AD (Byrd et al., 2017Byrd A.L. Deming C. Cassidy S.K.B. Harrison O.J. Ng W.I. Conlan S. et al.Staphylococcus aureus and Staphylococcus epidermidis strain diversity underlying pediatric atopic dermatitis.Sci Transl Med. 2017; 9eaal4651Crossref PubMed Scopus (244) Google Scholar; Totté et al., 2019Totté J.E.E. Pardo L.M. Fieten K.B. Vos M.C. van den Broek T.J. Schuren F.H.J. et al.Nasal and skin microbiome are associated with disease severity in pediatric atopic dermatitis.Br J Dermatol. 2019; 181: 796-804Crossref PubMed Scopus (17) Google Scholar). Mild and moderate patients in our study did not have significantly higher staphylococcal colonization load than control patients (Supplementary Table S2) but did show a stepwise increase in hemolytic activity of staphylococcal isolates (Supplementary Table S3). Our results build on previous studies to suggest that a more precise understanding of staphylococcal strains is necessary to understand this interaction. In this study, we provide strong preliminary data that individual isolate level differences in staphylococcal hemolytic potential may influence the severity of AD in a predominantly pediatric population. A major driver in staphylococcal virulence is the agr system, and research aimed at the identification of small molecule inhibitors of this system in efforts to diminish S. aureus virulence has emerged over the past decade (Salam and Quave, 2018Salam A.M. Quave C.L. Targeting virulence in Staphylococcus aureus by chemical inhibition of the accessory gene regulator system in vivo.mSphere. 2018; 3: e00500-e00517Crossref PubMed Scopus (44) Google Scholar). In some cases, these studies have progressed toward acute and inflammatory models of skin disease. For example, solanamide B was recently reported to suppress δ-toxin‒induced disease in a modified epicutaneous murine disease model of AD (Baldry et al., 2018Baldry M. Nakamura Y. Nakagawa S. Frees D. Matsue H. Núñez G. et al.Application of an agr-specific antivirulence compound as therapy for Staphylococcus aureus-induced inflammatory skin disease.J Infect Dis. 2018; 218: 1009-1013Crossref PubMed Scopus (18) Google Scholar). Future studies on the association demonstrated in this study may provide insight into potential therapeutics targeting staphylococcal virulence. Although our study design was selective for staphylococci, our methods did not differentiate between hemolysis caused by distinct coagulase-negative species (Supplementary Figure S4). S. epidermidis strains may be important drivers of the association noted in this study. In vivo interactions between S. epidermidis and S. aureus are complex and warrant future investigation (Fredheim et al., 2015Fredheim E.G.A. Flægstad T. Askarian F. Klingenberg C. Colonisation and interaction between S. epidermidis and S. aureus in the nose and throat of healthy adolescents.Eur J Clin Microbiol Infect Dis. 2015; 34: 123-129Crossref PubMed Scopus (15) Google Scholar; Iwase et al., 2010Iwase T. Uehara Y. Shinji H. Tajima A. Seo H. Takada K. et al.Staphylococcus epidermidis Esp inhibits Staphylococcus aureus biofilm formation and nasal colonization.Nature. 2010; 465: 346-349Crossref PubMed Scopus (587) Google Scholar). Although we noted a correlation between AD severity and staphylococcal hemolysis in this study, this study does not address the causation of this effect. Our study design did not control for all potential confounders such as immunosuppression, topical product use, or bathing practices, but we plan to address these factors in future studies. Topical steroid use may reduce the level of colonization with S. aureus on atopic skin (Nilsson et al., 1992Nilsson E.J. Henning C.G. Magnusson J. Topical corticosteroids and Staphylococcus aureus in atopic dermatitis.J Am Acad Dermatol. 1992; 27: 29-34Abstract Full Text PDF PubMed Scopus (171) Google Scholar), which may have influenced the density of colonization in some of our patients because we did not control for this factor. Clearly, the relationship between these factors is complex and warrants dedicated study. This was a single-site pilot study, and further research is required to define the generalizability of these results. We plan to explore the relative contribution of individual staphylococcal isolates in the severity of AD in future studies. Data are available within the article or its supplementary materials. Emily A. Gurnee: http://orcid.org/0000-0002-6450-7826 Mengqing Xu: http://orcid.org/0000-0002-0599-7888 Caitlin J. Risener: http://orcid.org/0000-0002-4824-6064 Kelly Lehman: http://orcid.org/0000-0001-8946-0342 Kate Nelson: http://orcid.org/0000-0001-8783-2157 Robert A. Swerlick: http://orcid.org/0000-0002-9802-4144 Cassandra L. Quave: http://orcid.org/0000-0001-9615-7886 The authors state no conflict of interest. We acknowledge the Department of Dermatology Emory University (Atlanta, GA) development funds for financial support. This work has been performed in Atlanta, Georgia. Conceptualization: CLQ, RAS; Data Curation: EAG, CLQ; Formal Analysis: CLQ; Investigation: EAG, MX, CJR, KL, KN; Methodology: CLQ, EAG, RAS; Project Administration: CLQ; Resources: CLQ, EAG; Supervision: CLQ, EAG; Validation: CLQ, EAG; Visualization: CLQ, EAG; Writing - Original Draft Preparation: EAG; Writing - Review and Editing: EAG, MX, CJR, KL, KN, RAS, CLQ Download .pdf (1.94 MB) Help with pdf files Supplementary Data