Viral DNA polymerase structures reveal mechanisms of antiviral drug resistance.

DNA polymerases are important drug targets, and many structural studies have captured them in distinct conformations. However, a detailed understanding of the impact of polymerase conformational dynamics on drug resistance is lacking. We determined cryoelectron microscopy (cryo-EM) structures of DNA-bound herpes simplex virus polymerase holoenzyme in multiple conformations and interacting with antivirals in clinical use. These structures reveal how the catalytic subunit Pol and the processivity factor UL42 bind DNA to promote processive DNA synthesis. Unexpectedly, in the absence of an incoming nucleotide, we observed Pol in multiple conformations with the closed state sampled by the fingers domain. Drug-bound structures reveal how antivirals may selectively bind enzymes that more readily adopt the closed conformation. Molecular dynamics simulations and the cryo-EM structure of a drug-resistant mutant indicate that some resistance mutations modulate conformational dynamics rather than directly impacting drug binding, thus clarifying mechanisms that drive drug selectivity.

The immunobiology of corneal HSV-1 infection and herpetic stromal keratitis.

SUMMARYHuman alphaherpesvirus 1 (HSV-1) is a highly successful neurotropic pathogen that primarily infects the epithelial cells lining the orofacial mucosa. After primary lytic replication in the oral, ocular, and nasal mucosal epithelial cells, HSV-1 establishes life-long latency in neurons within the trigeminal ganglion. Patients with compromised immune systems experience frequent reactivation of HSV-1 from latency, leading to virus entry in the sensory neurons, followed by anterograde transport and lytic replication at the innervated mucosal epithelial surface. Although recurrent infection of the corneal mucosal surface is rare, it can result in a chronic immuno-inflammatory condition called herpetic stromal keratitis (HSK). HSK leads to gradual vision loss and can cause permanent blindness in severe untreated cases. Currently, there is no cure or successful vaccine to prevent latent or recurrent HSV-1 infections, posing a significant clinical challenge to managing HSK and preventing vision loss. The conventional clinical management of HSK primarily relies on anti-virals to suppress HSV-1 replication, anti-inflammatory drugs (such as corticosteroids) to provide symptomatic relief from pain and inflammation, and surgical interventions in more severe cases to replace damaged cornea. However, each clinical treatment strategy has limitations, such as local and systemic drug toxicities and the emergence of anti-viral-resistant HSV-1 strains. In this review, we summarize the factors and immune cells involved in HSK pathogenesis and highlight alternate therapeutic strategies for successful clinical management of HSK. We also discuss the therapeutic potential of immunoregulatory cytokines and immunometabolism modulators as promising HSK therapies against emerging anti-viral-resistant HSV-1 strains.

Mucosal Site Detection of Herpes Simplex Virus in Neonates.

From 2001 to 2023, 17 (14%) of 120 neonates with confirmed herpes simplex virus infection tested positive for herpes simplex virus by polymerase chain reaction (PCR) from only mucosal sites without a clinical mucosal lesion. Whether mucosal PCR positivity reflects early infection that may lead to recognizable disease, transient colonization, or a false-positive PCR result remains a clinical conundrum and warrants further study.

Israeli neonatal herpes simplex infection: Unique epidemiology and clinical profile.

To gather national level data on Israeli neonatal HSV (NHSV) infection and to evaluate the distinct clinical characteristics of NHSV and neonatal enteroviral meningitis (NEM). Israeli NHSV patients, hospitalized between January 2015 and April 2022 in 22 medical centers were assessed, together with NEM patients, hospitalized at Sheba Medical Center during the same period. NHSV demographic and clinical characteristics were documented and compared to those of NEM. Eighty-five NHSV (73% males) and 130 NEM (62% males) patients were included. The incidence of NHSV was 5.9/100 000 live births, the common phenotype and HSV type were SEM (53%) and HSV1 (91%), respectively. Horizontal transmission was suspected in 50% cases (of which 67% underwent a Jewish ritual circumcision with direct wound sucking, 33% had relatives with highly suspicious herpetic lesions). Compared with NEM, NHSV tends to present with rash (14% vs. 60%, p-value < 0.01) and seizures (0% vs. 6%, p-value 0.02), while fever, irritability and poor feeding appear more frequently in NEM (94% vs. 18%, p-value < 0.01; 37% vs. 1%, p-value < 0.01; 25% vs. 1%, p-value < 0.01 respectively). Of NEM patients, 28% were treated with acyclovir. Our results mark a decrease in the incidence rate of NHSV in Israel and a prominent mode of horizontal infection acquisition. We underscore the unique localized phenotype of NHSV, in contrast to enterovirus, which tends to cause a systemic disease with constitutional symptoms. These findings should be considered when evaluating the need for comprehensive empirical treatment for HSV in the context of neonatal fever, or according to a certain clinical presentation.

Development of pH-responsive Hydrogel from Copolymers of Artemisia vulgaris Seed Mucilage, Mucin, and poly(methacrylate) for Controlled Delivery of Acyclovir Sodium.

To cope with the constraints of conventional drug delivery systems, site-specific drug delivery systems are the major focus of researchers. The present research developed water-swellable, pH-responsive methacrylic acid-based hydrogel scaffolds of Artemisia vulgaris seed mucilage with mucin and loaded with acyclovir sodium as a model drug. The developed hydrogel discs are evaluated for diverse parameters. Drug loading efficiency in all formulations ranges from 63% to 75%. The hydrogels exhibited pH-dependent swelling, displaying optimum swelling in a phosphate buffer (pH 7.4), and insignificant swelling in an acidic buffer (pH 1.2), in addition, they responded well to electrolyte concentrations. The sol-gel fraction is estimated ranging from 60 to 95%. Dissolution studies unveiled sustained drug release for 24 h in a phosphate buffer of pH 7.4, exhibiting zero-order release kinetics. Moreover, FTIR spectra confirmed the drug-excipient compatibility. SEM photomicrographs revealed a rough and porous surface of hydrogel discs with several pores and channels. The PXRD diffractograms exposed the amorphous nature of the polymeric blends. The findings of acute toxicity studies proved the developed hydrogel network is biocompatible. Therefore, these outcomes connote the newly created network as a smart delivery system, able to dispatch acyclovir sodium into the intestinal segment for a prolonged period.

Varicella-Zoster Virus Reactivation After Pediatric Allogeneic Hematopoietic Stem Cell Transplantation, Single-Center Experience of Acyclovir Prophylaxis.

BACKGROUND: Varicella-zoster virus (VZV) reactivation is the most common infectious complication in the late posthematopoietic stem cell transplantation (HSCT) period and is reported as 16%-41%. Acyclovir prophylaxis is recommended for at least 1 year after HSCT to prevent VZV infections. However, studies on the most appropriate prophylaxis are ongoing in pediatric patients. METHODS: Patients who underwent allogeneic HSCT between January 1, 1996 and January 1, 2020 were retrospectively analyzed to outline the characteristics of VZV reactivation after allogeneic HSCT in pediatric patients using 6 months acyclovir prophylaxis. RESULTS: There were 260 patients and 273 HSCTs. Median age was 10.43 (0.47-18.38), and 56% was male. Median follow-up was 2325 days (18-7579 days). VZV reactivation occurred in 21.2% (n = 58) at a median of 354 (55-3433) days post-HSCT. The peak incidence was 6-12 months post-HSCT (43.1%). Older age at HSCT, female gender, history of varicella infection, lack of varicella vaccination, low lymphocyte, CD4 count, and CD4/CD8 ratio at 9 and 12 months post-HSCT was found as a significant risk for herpes zoster (HZ) in univariate analysis, whereas history of varicella infection and low CD4/CD8 ratio at 12 months post-HSCT was an independent risk factor in multivariate analysis. CONCLUSIONS: Tailoring acyclovir prophylaxis according to pre-HCT varicella history, posttransplant CD4 T lymphocyte counts and functions, and ongoing immunosuppression may help to reduce HZ-related morbidity and mortality.

Efficacy of photobiomodulation therapy in recurrent herpes labialis management: a randomized controlled trial.

OBJECTIVES: This study is aimed at assessing the therapeutic efficacy of photobiomodulation therapy (PBMT) for the management of recurrent herpes labialis (RHL) by evaluating both pain and clinical recovery. MATERIAL AND METHODS: A randomized, double-blind, controlled trial was conducted on 40 patients with RHL, and they were randomly divided into two groups, where 20 patients received treatment with PBMT (650 nm, 100 mW, 4.7 J/cm2), continuous mode, for 120 s, and placebo cream, while another 20 patients (control group) were treated with acyclovir cream 5% (5 times/5 days) and passive laser. Pain was assessed at five different times. The day when the complete disappearance of the pain was observed and the day when the crust fell off spontaneously were also recorded. RESULTS: The pain level in the control group was significantly higher than that in the PBMT group after the second application of the laser, while the differences were not significant between the two groups at other times. The pain in the PBMT group disappeared faster than that in the control group, but the difference was not significant in terms of clinical recovery. CONCLUSIONS: Photobiomodulation therapy of herpes labialis reduced pain significantly faster than acyclovir, but there was no difference in healing time between the groups in light of the parameters used in this study. CLINICAL RELEVANCE: PBMT is a promising treatment that may be an effective alternative to acyclovir in the management of recurrent herpes labialis. TRIAL REGISTRATION ISRCTN: com ID: ISRCTN87606522.

First Acyclovir Determination Procedure via Electrochemically Activated Screen-Printed Carbon Electrode Coupled with Well-Conductive Base Electrolyte.

In this work, a new voltammetric procedure for acyclovir (ACY) trace-level determination has been described. For this purpose, an electrochemically activated screen-printed carbon electrode (aSPCE) coupled with well-conductive electrolyte (CH3COONH4, CH3COOH and NH4Cl) was used for the first time. A commercially available SPCE sensor was electrochemically activated by conducting cyclic voltammetry (CV) scans in 0.1 mol L-1 NaOH solution and rinsed with deionized water before a series of measurements were taken. This treatment reduced the charge transfer resistance, increased the electrode active surface area and improved the kinetics of the electron transfer. The activation step and high conductivity of supporting electrolyte significantly improved the sensitivity of the procedure. The newly developed differential-pulse adsorptive stripping voltammetry (DPAdSV) procedure is characterized by having the lowest limit of detection among all voltammetric procedures currently described in the literature (0.12 nmol L-1), a wide linear range of the calibration curve (0.5-50.0 and 50.0-1000.0 nmol L-1) as well as extremely high sensitivity (90.24 nA nmol L-1) and was successfully applied in the determination of acyclovir in commercially available pharmaceuticals.

Development and Validation of an HPLC-UV Method for the Quantification of Acyclovir and Ganciclovir in the Plasma of Pediatric Immunocompromised Patients.

Acyclovir and ganciclovir comprise the prophylaxis and treatment of herpesvirus and cytomegalovirus infections occurring in immunocompromised patients. Their therapeutic drug monitoring is fundamental because of interindividual variability leading to side effects and drug resistance and is performed through several techniques, such as liquid chromatography coupled with UV spectrophotometry (HPLC-UV) or mass spectrometry (LC-MS/MS). Therefore, we developed and validated a low-cost, non-time-consuming, and low-sample-consuming HPLC-UV method. Briefly, 100 µL of sample was used for sample preparation, mainly consisting of precipitation through organic solvent. In total, 20 µL was injected into the instrument. Chromatographic separation was obtained eluting mobile phases A (10 mM ammonium formiate 0.01% formic acid) and B (acetonitrile) on a Poroshell 120 SB-C8 2.1 × 150 mm, 2.7 µm for 12 min isocratically (97:3; A:B) at a flow rate of 0.2 mL/min. The linearity range (0.5-40 mg/L) of the method allowed us to quantify both the Cmin and Cmax of acyclovir and ganciclovir. Plasma concentrations measured on a small cohort of patients undergoing acyclovir (31) and ganciclovir (9) treatment by the proposed method and the LC-MS/MS methods, already in use, were significantly correlated. The proposed HPLC-UV method may be implemented in diagnostics as an alternative method in case of the unavailability of the LC-MS/MS system.

Degradation Acyclovir Using Sodium Hypochlorite: Focus on Byproducts Analysis, Optimal Conditions and Wastewater Application.

In recent years, the environmental impact of pharmaceutical residues has emerged as a pressing global concern, catalyzed by their widespread usage and persistence in aquatic ecosystems. Among these pharmaceuticals, acyclovir (ACV) stands out due to its extensive prescription during medical treatments for herpes simplex virus, chickenpox, and shingles, as well as its heightened usage amidst the COVID-19 pandemic. ACV is excreted largely unchanged by the human body, leading to significant environmental release through wastewater effluents. The urgency of addressing ACV's environmental impact lies in its potential to persist in water bodies and affect aquatic life. This persistence underscores the critical need for effective degradation strategies that can mitigate its presence in aquatic systems. This study focuses on employing sodium hypochlorite as an oxidative agent for the degradation of ACV, leveraging its common use in wastewater treatment plants. Our research aims to explore the kinetics of ACV degradation, identify and characterize its degradation byproducts, and optimize the conditions under which complete degradation can be achieved. By assessing the efficiency of sodium hypochlorite in real wastewater samples, this study seeks to provide practical insights into mitigating ACV contamination in aquatic environments. The novelty of this research lies in its comprehensive approach to understanding the degradation pathways of ACV and evaluating the feasibility of using sodium hypochlorite as a sustainable solution in wastewater treatment. By addressing the environmental concerns associated with ACV and offering practical solutions, this study contributes to the broader goal of sustainable pharmaceutical waste management and environmental stewardship.